Topical compositions

ABSTRACT

Provided herein are anhydrous compositions that include at least one viscosity increasing agent, at least one organic solvent and at least one humectant. Such compositions may also include at least one active pharmaceutical ingredient (API) and/or at least one water repellant. Related compositions, methods and kits are also provided.

RELATED APPLICATION DATA

This application claims the benefit of U.S. Provisional PatentApplication Ser. No. 61/504,628, filed Jul. 5, 2011, and U.S.Provisional Patent Application Ser. No. 61/610,137, filed Mar. 13, 2012,the disclosure of each of which are incorporated herein by reference intheir entirety.

FIELD OF THE INVENTION

The present invention is directed to pharmaceutical compositions. Inparticular, the present invention is directed to anhydrous topicalpharmaceutical compositions.

BACKGROUND OF THE INVENTION

Acne vulgaris is the most common skin disease in the United States. Itis estimated that 40 to 50 million Americans have acne, including 80% ofpeople between the ages of 11 and 30. The annual direct costs associatedwith the treatment of acne exceeded $2.8 billion in 2007, with themajority of those costs attributable to prescription drugs. In addition,acne causes both physical and psychological effects, including permanentscarring, anxiety, depression, and poor self-esteem. Even in cases ofmild acne, the social stigma associated with the disease frequentlyresults in significant emotional distress and other psychologicalissues. Due to its social impact, frequency of recurrence of relapse,and necessary maintenance over a prolonged course of therapy, theAmerican Academy of Dermatologists have recommend that acne vulgaris bere-classified and investigated as a chronic disease.

Acne vulgaris results from the complex interplay of four majorpathogenic factors: 1) overproduction of sebum by the sebaceous gland;2) abnormal keratinization in the follicle; 3) colonization of the hairfollicles by the anaerobic, lipophilic bacterium Propionibacteriumacnes, or P. acnes; and 4) release of inflammatory mediators into theskin. All acne lesions begin when the combination of excess sebum andabnormal epithelial desquamation clog up a follicle, forming amicroscopic lesion known as a microcomedo. The anaerobic, lipid-richenvironment of the microcomedo provides an ideal location for P. acnesproliferation. Each microcomedo may progress to form a non-inflammatoryopen or closed comedone (commonly referred to as a “blackhead” or“whitehead,” respectively), or an inflammatory lesion that may befurther categorized as a papule, pustule, nodule, or cyst.

The complexity of the disease may require multiple treatments that mayspan oral and topical antimicrobials, oral and topical retinoids, oralcontraceptives and other prescription skin cleansers. The most effectivetherapies for acne are those that may safely address more than one ofthe major causes of acne pathogenesis.

Antibiotics were the first successful acne treatment due to theirantimicrobial and anti-inflammatory properties. Both topical andsystemic antibiotics have been very successful, but the protractedtreatment periods required have led to the development of resistance ofP. acnes and in other non-targeted (and potentially pathogenic)commensal organisms. Combining antibiotics with topical retinoidstargets three of the four major pathogenic factors associated with acne(all but sebum production). The oral retinoid isotretinoin (e.g.,Accutane®) is the only drug-implicated to affect all four pathogenicfactors associated with acne. However, the severity of its potentialside effects (known teratogen and linked to depression, psychosis andsuicide) has limited its use and led to numerous lawsuits.

While the problems associated with isotretinoin are the most severe, allof the current acne medications have some adverse effects. The majorityof topical treatments lead to dryness, irritation and peeling of theskin, and oral antibiotics may cause gastrointestinal tract irritation,photosensitivity of skin, headache, dizziness, anemia, bone and jointpain, nausea and/or depression.

The most commonly prescribed drugs from acne are antibiotics, includingbenzoyl peroxide, clindamycin and erythromycin, either alone or incombination, and retinoids, including adapalene, tretinoin andtazarotene, either alone or in combination with an antibiotic.Treatments may include combination drugs or combination therapies. Forexample, a retinoid may be prescribed for application in the morning andan antibiotic for application in the evening. Each of these commonlyprescribed drugs, however, has disadvantages that often reduce theeffectiveness of the therapy.

For example, benzoyl peroxide may be the most effective topical medicinefor acne and may result in a rapid reduction in P. acnes. It also doesnot induce drug resistance in the P. acnes and, when combined with otherantibiotics, may reduce the rate at which drug resistance develops.However, benzoyl peroxide commonly results in irritation and dryness ofthe skin and bleaches fabric. Additionally, about 2% of patients have anallergic reaction to benzoyl peroxide.

Clindamycin and erythromycin as monotherapies may be limited ineffectiveness because of the development of drug resistant strains of P.acnes. In fact, it is estimated that up to 75% of P. acnes is alreadyresistant to these antibiotics. However, these products are available ina number of different bases and are typically non-irritating andnon-staining.

Retinoids mainly target comedonal acne but may also reduce inflammatorylesions. Retinoids, however, are slow to produce visible results and mayproduce irritation, redness and peeling. In light of these limitationsof retinoids, they are typically not used alone.

Combinations of these various drugs are also available. For example,clindamycin and benzoyl peroxide combination drugs are available as wellas combinations of erythromycin and benzoyl peroxide, combinations ofadapalene and benzoyl peroxide and combinations of clindamycin andtretinoin. These drugs may have improved efficacy over the individualdrugs but also carry with them the limitations of their constituents.Furthermore, none of these combinations address all four of the causesof acne.

In addition to the limitations of the available drugs themselves, anumber of additional factors may affect compliance with a treatmentregime and, therefore, may reduce the regime's overall efficacy. Thesefactors include the degree of irritation of the product, the time tonoticeable results, the aesthetics of the product, the repeatabilityrequired, the effect on clothing and other items the product comes incontact with and the convenience of the packaging and storage of theproduct. If the product irritates the skin, the patient may discontinueuse. If the product takes too long to provide visible results, thepatient may get frustrated and stop treatment. If the product feelsgreasy, leaves a visible residue or is powdery and grating, the patientmay be less likely to maintain the regime. If the product stains orbleaches clothes, bedding or other fabrics, the patient may discontinueits use. Finally, if the product is too difficult to use or store, thepatient may be unlikely to use it or to follow the use and/or storagedirections.

It may be difficult to obtain compositions that address some or all ofthese factors. It may be even more difficult to obtain anhydrouscompositions that address some or all of these factors. As such, newcompositions and methods for making such compositions may be desirable.

SUMMARY OF THE INVENTION

Provided according to some embodiments of the invention are anhydroustopical compositions that include at least one viscosity increasingagent, at least one organic solvent and at least one humectant. In someembodiments, the at least one viscosity increasing agent is present inthe composition at a concentration in a range of 0.5 to 30 weightpercent; the at least one organic solvent is present in the compositionat a concentration in a range of 50 to 90 weight percent; and the atleast one humectant is present in the composition at a concentration ina range of 2 to 20 weight percent.

In some embodiments of the invention, the topical anhydrous compositionsfurther include at least one water repellant. In some embodiments, theat least one viscosity increasing agent is present in the composition ata concentration in a range of 0.5 to 30 weight percent; the at least oneorganic solvent is present in the composition at a concentration in arange of 50 to 90 weight percent; the at least one humectant is presentin the composition at a concentration in a range of 2 to 20 weightpercent; and the at least one water repellent is present in thecomposition at a concentration in a range of 0.5 to 15 weight percent.

In some embodiments, the topical anhydrous compositions further includea water reactive active pharmaceutical ingredient (API). In someembodiments, the water reactive API releases nitric oxide (NO) uponcontact with water. In some embodiments, the water reactive API includesa diazeniumdiolate functional group. In some embodiments, the waterreactive API includes a NO-releasing co-condensed silica.

Provided also in some embodiments are packaged nitric oxide-releasingtopical gels having a shelf life of at least four weeks. Furtherprovided are packaged nitric oxide-releasing topical gels having usefullife of at least 30 days and in some embodiments at least 60 days.

A further aspect of the present invention comprises a method of treatinga dermatological condition comprising applying an anhydrous topicalcomposition of the present invention to the skin of a patient to treatthe dermatological condition. In some embodiments, an anhydrous topicalcomposition comprises at least one API. In other embodiments, ananhydrous topical composition comprises a vehicle and thus does notinclude at least one API. In certain embodiments, an anhydrous topicalcomposition of the present invention is applied to the skin of a patientto treat acne.

BRIEF DESCRIPTION OF THE DRAWINGS

The following drawings are provided to illustrate various aspects of thepresent inventive concept and are not intended to limit the scope of thepresent invention unless specified herein.

FIG. 1 is a process flow diagram illustrating manufacture and packagingof a topical therapeutic according to some embodiments of the presentinventive concept.

FIG. 2 is a flowchart illustrating operations/steps in a process formanufacturing a topical therapeutic according to some embodiments of thepresent inventive concept.

FIG. 3 is a flowchart illustrating further operations/steps in a processfor manufacturing a topical therapeutic according to some embodiments ofthe present inventive concept

FIGS. 4A through 4C are drawings of a rotor-stator homogenizer accordingto some embodiments of the present inventive concept.

FIG. 5 shows the reduction in P. acnes for a control blank, a gelvehicle and a composition according to an embodiment of the invention.

FIG. 6 shows the nitric oxide release detected via chemiluminescence inreal time from a 2% API loaded anhydrous gel, wherein the API is anitric oxide releasing macromolecule, when placed in phosphate bufferedsaline at a representative skin pH of 6.0. Additionally, the totalnitric oxide release from the anhydrous gel in nmol NO/mg gel is shown,demonstrating complete release in under 30 minutes.

FIG. 7 shows the stability of a 2% API loaded anhydrous gel, wherein theAPI is a nitric oxide releasing macromolecule, packaged in various sizedaluminum tubes and maintained in the following conditions: refrigerated(4° C.), room temperature with 75% relative humidity, and 40° C. with75% relative humidity. Percent NO recovery values were measured by thetotal NO release detected via chemiluminescence at each time point andpresented as a percentage of the nitric oxide loading at time=0.

FIG. 8 shows a graph of nitric oxide recovery versus time for a 2%anhydrous gel in 2 gram tubes.

DETAILED DESCRIPTION OF EMBODIMENTS OF THE INVENTION

The foregoing and other aspects of the present invention will now bedescribed in more detail with respect to the description andmethodologies provided herein. It should be appreciated that theinvention may be embodied in different forms and should not be construedas limited to the embodiments set forth herein. Rather, theseembodiments are provided so that this disclosure will be thorough andcomplete, and will fully convey the scope of the invention to thoseskilled in the art.

The terminology used in the description of the invention herein is forthe purpose of describing particular embodiments only and is notintended to be limiting of the invention. As used in the description ofthe embodiments of the invention and the appended claims, the singularforms “a,” “an” and “the” are intended to include the plural forms aswell, unless the context clearly indicates otherwise. Also, as usedherein, “and/or” refers to and encompasses any and all possiblecombinations of one or more of the associated listed items. Furthermore,the term “about,” as used herein when referring to a measurable valuesuch as an amount of a compound, dose, time, temperature, and the like,is meant to encompass variations of 20%, 10%, 5%, 1%, 0.5%, or even 0.1%of the specified amount. When a range is employed (e.g., a range from xto y) it is it meant that the measurable value is a range from about xto about y, or any range therein, such as about x₁ to about y₁, etc. Itwill be further understood that the terms “comprises” and/or“comprising,” when used in this specification, specify the presence ofstated features, integers, steps, operations, elements, and/orcomponents, but do not preclude the presence or addition of one or moreother features, integers, steps, operations, elements, components,and/or groups thereof. Unless otherwise defined, all terms, includingtechnical and scientific terms used in the description, have the samemeaning as commonly understood by one of ordinary skill in the art towhich this invention belongs.

All patents, patent applications and publications referred to herein areincorporated by reference in their entirety. In the event of conflictingterminology, the present specification is controlling.

The embodiments described in one aspect of the present invention are notlimited to the aspect described. The embodiments may also be applied toa different aspect of the invention as long as the embodiments do notprevent these aspects of the invention from operating for its intendedpurpose.

CHEMICAL DEFINITIONS

As used herein the term “alkyl” refers to C1-20 inclusive, linear (i.e.,“straight-chain”), branched, or cyclic, saturated or at least partiallyand in some cases fully unsaturated (i.e., alkenyl and alkynyl)hydrocarbon chains, including for example, methyl, ethyl, propyl,isopropyl, butyl, isobutyl, tert-butyl, pentyl, hexyl, octyl, ethenyl,propenyl, butenyl, pentenyl, hexenyl, octenyl, butadienyl, propynyl,butynyl, pentynyl, hexynyl, heptynyl, and allenyl groups. “Branched”refers to an alkyl group in which a lower alkyl group, such as methyl,ethyl or propyl, is attached to a linear alkyl chain. Exemplary branchedalkyl groups include, but are not limited to, isopropyl, isobutyl,tert-butyl. “Lower alkyl” refers to an alkyl group having 1 to 8 carbonatoms (i.e., a C1-8 alkyl), e.g., 1, 2, 3, 4, 5, 6, 7, or 8 carbonatoms. “Higher alkyl” refers to an alkyl group having 8 to 20 or morecarbon atoms, e.g., 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, ormore carbon atoms. In certain embodiments, “alkyl” refers, inparticular, to C1-5 straight-chain alkyls. In other embodiments, “alkyl”refers, in particular, to C1-5 branched-chain alkyls.

Alkyl groups may optionally be substituted (a “substituted alkyl”) withone or more alkyl group substituents, which may be the same ordifferent. The term “alkyl group substituent” includes but is notlimited to alkyl, substituted alkyl, halo, arylamino, acyl, hydroxyl,aryloxyl, alkoxyl, alkylthio, arylthio, aralkyloxyl, aralkylthio,carboxyl, alkoxycarbonyl, oxo, and cycloalkyl. There may be optionallyinserted along the alkyl chain one or more oxygen, sulfur or substitutedor unsubstituted nitrogen atoms, wherein the nitrogen substituent ishydrogen, lower alkyl (also referred to herein as “alkylaminoalkyl”), oraryl.

Thus, as used herein, the term “substituted alkyl” includes alkylgroups, as defined herein, in which one or more atoms or functionalgroups of the alkyl group are replaced with another atom or functionalgroup, including for example, alkyl, substituted alkyl, halogen, aryl,substituted aryl, alkoxyl, hydroxyl, nitro, amino, alkylamino,dialkylamino, sulfate, and mercapto.

The term “aryl” is used herein to refer to an aromatic substituent thatmay be a single aromatic ring, or multiple aromatic rings that are fusedtogether, linked covalently, or linked to a common group, such as, butnot limited to, a methylene or ethylene moiety. The common linking groupalso may be a carbonyl, as in benzophenone, or oxygen, as indiphenylether, or nitrogen, as in diphenylamine. The term “aryl”specifically encompasses heterocyclic aromatic compounds. The aromaticring(s) may comprise phenyl, naphthyl, biphenyl, diphenylether,diphenylamine and benzophenone, among others. In particular embodiments,the term “aryl” means a cyclic aromatic comprising 5 to 10 carbon atoms,e.g., 5, 6, 7, 8, 9, or 10 carbon atoms, and including 5- and 6-memberedhydrocarbon and heterocyclic aromatic rings.

The aryl group may be optionally substituted (a “substituted aryl”) withone or more aryl group substituents, which may be the same or different,wherein “aryl group substituent” includes alkyl, substituted alkyl,aryl, substituted aryl, aralkyl, hydroxyl, alkoxyl, aryloxyl,aralkyloxyl, carboxyl, acyl, halo, nitro, alkoxycarbonyl,aryloxycarbonyl, aralkoxycarbonyl, acyloxyl, acylamino, aroylamino,carbamoyl, alkylcarbamoyl, dialkylcarbamoyl, arylthio, alkylthio,alkylene, and —NR1R″, wherein R1 and R″ may each be independentlyhydrogen, alkyl, substituted alkyl, aryl, substituted aryl, and aralkyl.

Thus, as used herein, the term “substituted aryl” includes aryl groups,as defined herein, in which one or more atoms or functional groups ofthe aryl group are replaced with another atom or functional group,including for example, alkyl, substituted alkyl, halogen, aryl,substituted aryl, alkoxyl, hydroxyl, nitro, amino, alkylamino,dialkylamino, sulfate, and mercapto. Specific examples of aryl groupsinclude, but are not limited to, cyclopentadienyl, phenyl, furan,thiophene, pyrrole, pyran, pyridine, imidazole, benzimidazole,isothiazole, isoxazole, pyrazole, pyrazine, triazine, pyrimidine,quinoline, isoquinoline, indole, carbazole, and the like.

“Cyclic” and “cycloalkyl” refer to a non-aromatic mono- or multicyclicring system of 3 to 10 or more carbon atoms, e.g., 3, 4, 5, 6, 7, 8, 9,10, or more carbon atoms. The cycloalkyl group may be optionallypartially unsaturated. The cycloalkyl group also may be optionallysubstituted with an alkyl group substituent as defined herein, oxo,and/or alkylene. There may be optionally inserted along the cyclic alkylchain one or more oxygen, sulfur or substituted or unsubstitutednitrogen atoms, wherein the nitrogen substituent is hydrogen, alkyl,substituted alkyl, aryl, or substituted aryl, thus providing aheterocyclic group. Representative monocyclic cycloalkyl rings includecyclopentyl, cyclohexyl, and cycloheptyl. Multicyclic cycloalkyl ringsinclude adamantyl, octahydronaphthyl, decalin, camphor, camphane, andnoradamantyl.

“Alkoxyl” refers to an alkyl-O— group wherein alkyl is as previouslydescribed. The term “alkoxyl” as used herein may refer to, for example,methoxyl, ethoxyl, propoxyl, isopropoxyl, butoxyl, f-butoxyl, andpentoxyl. The term “oxyalkyl” may be used interchangeably with“alkoxyl”. In some embodiments, the alkoxyl has 1, 2, 3, 4, or 5carbons.

“Aralkyl” refers to an aryl-alkyl group wherein aryl and alkyl are aspreviously described, and included substituted aryl and substitutedalkyl. Exemplary aralkyl groups include benzyl, phenylethyl, andnaphthylmethyl.

“Alkylene” refers to a straight or branched bivalent aliphatichydrocarbon group having from 1 to 20 or more carbon atoms, e.g., 1, 2,3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or morecarbon atoms. The alkylene group may be straight, branched or cyclic.The alkylene group also may be optionally unsaturated and/or substitutedwith one or more “alkyl group substituents.” There may be optionallyinserted along the alkylene group one or more oxygen, sulfur orsubstituted or unsubstituted nitrogen atoms (also referred to herein as“alkylaminoalkyl”), wherein the nitrogen substituent is alkyl aspreviously described. Exemplary alkylene groups include methylene(—CH2-); ethylene (—CH2-CH2-); propylene (—(CH2)3-); cyclohexylene(—C6H10-); —CH═CH—CH═CH—; —CH═CH—CH2-; wherein each of q and r isindependently an integer from 0 to 20, e.g., 0, 1, 2, 3, 4, 5, 6, 7, 8,9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20, or any range therein,and R is hydrogen or lower alkyl; methylenedioxyl (—O—CH2-O—); andethylenedioxyl (—O—(CH2)2-O—). An alkylene group may have 2 to 3 carbonatoms and may further have 6-20 carbons.

“Arylene” refers to a bivalent aryl group. An exemplary arylene isphenylene, which may have ring carbon atoms available for bonding inortho, meta, or para positions with regard to each other, i.e.,respectively. The arylene group may also be napthylene. The arylenegroup may be optionally substituted (a “substituted arylene”) with oneor more “aryl group substituents” as defined herein, which may be thesame or different.

“Aralkylene” refers to a bivalent group that contains both alkyl andaryl groups. For example, aralkylene groups may have two alkyl groupsand an aryl group (i.e., -alkyl-aryl-alkyl-), one alkyl group and onearyl group (i.e., -alkyl-aryl-) or two aryl groups and one alkyl group(i.e., -aryl-alkyl-aryl-).

The term “amino” and “amine” refer to nitrogen-containing groups such asNR3, NH3, NHR2, and NH2R, wherein R may be alkyl, branched alkyl,cycloalkyl, aryl, alkylene, arylene, aralkylene. Thus, “amino” as usedherein may refer to a primary amine, a secondary amine, or a tertiaryamine. In some embodiments, one R of an amino group may be a cationstabilized diazeniumdiolate (i.e., NONO—X+).

The terms “cationic amine” and “quaternary amine” refer to an aminogroup having an additional (i.e., a fourth) group, for example ahydrogen or an alkyl group bonded to the nitrogen. Thus, cationic andquartemary amines carry a positive charge.

The term “alkylamine” refers to the -alkyl-NH2 group.

The term “carbonyl” refers to the —(C═O)— group.

The term “carboxyl” refers to the —COOH group and the term “carboxylate”refers to an anion formed from a carboxyl group, i.e., —COO—.

The terms “halo”, “halide”, or “halogen” as used herein refer to fluoro,chloro, bromo, and iodo groups.

The term “hydroxyl” and “hydroxy” refer to the —OH group.

The term “hydroxyalkyl” refers to an alkyl group substituted with an —OHgroup.

The term “mercapto” or “thio” refers to the —SH group. The term “silyl”refers to groups comprising silicon atoms (Si).

As used herein the term “alkoxysilane” refers to a compound comprisingone, two, three, or four alkoxy groups bonded to a silicon atom. Forexample, tetraalkoxysilane refers to Si(OR)4, wherein R is alkyl. Eachalkyl group may be the same or different. An “alkylsilane” refers to analkoxysilane wherein one or more of the alkoxy groups has been replacedwith an alkyl group. Thus, an alkylsilane comprises at least onealkyl-Si bond. The term “fluorinated silane” refers to an alkylsilanewherein one of the alkyl groups is substituted with one or more fluorineatoms. The term “cationic or anionic silane” refers to an alkylsilanewherein one of the alkyl groups is further substituted with an alkylsubstituent that has a positive (i.e., cationic) or a negative (i.e.anionic) charge, or may become charged (i.e., is ionizable) in aparticular environment (i.e., in vivo).

The term “silanol” refers to a Si—OH group.

Pharmaceutical Compositions

One aspect of the present invention is directed to pharmaceuticalcompositions. Provided according to some embodiments of the inventionare pharmaceutical compositions that include at least one activepharmaceutical ingredient (API). According to other embodiments of thepresent invention, provided are pharmaceutical compositions that do notinclude at least one API. Compositions described herein that do not havean API are referred to herein as “vehicles.” Pharmaceutical compositionsaccording to embodiments of the invention may, therefore, include atleast one API or may be vehicles that do not include at least one API.In some embodiments, the pharmaceutical compositions are anhydrous, andin some embodiments, the pharmaceutical compositions are topicalcompositions. “Anhydrous,” as used herein means that there is no directaddition of water to a composition of the present invention. However,those skilled in the art will recognize that water may be physicallyand/or chemically absorbed by a composition and/or by one or moreingredients in a composition at any time during the preparation,storage, and/or use of a composition of the present invention (i.e.,indirect addition of water to the composition). In some embodiments, theterm “anhydrous” means that a composition has a water content of lessthan 5% by weight of the composition or any range therein. A compositionof the present invention may have a water content of less than 5, 4.5,4, 3.5, 3, 2.5, 2, 1.5, 1, 0.5%, or any range therein, by weight of thecomposition. Water content may be measured by methods known to those ofskill in the art, such as, but not limited to, Karl Fischer titration.

In some embodiments of the invention, the pharmaceutical composition isa topical composition, which may also be referred to as a“dermatological composition” herein. A topical composition may beapplied to body surfaces, including skin, mucous membranes, scalp, hairand/or nails.

Provided according to embodiments of the invention are anhydrouspharmaceutical compositions that include excipients that include atleast one viscosity agent, at least one solvent and at least onehumectant. The term excipient refers to “inert” constituents ofpharmaceutically acceptable compositions. The term “inert” indicatesthat such constituents are not considered active pharmaceuticalingredients, such as a nitric oxide-releasing compound or otherantimicrobial compounds, anti-inflammatory agents, pain-relievers,immunosuppressants and vasodilators. However, as one of ordinary skillin the art will understand, the excipients may provide beneficial ortherapeutic action to the skin (e.g., moisturize, provideanti-inflammatory effects) that may directly affect a topical ailment.The excipients may also indirectly affect the treatment of topicalailments by affecting the stability of NO-releasing compounds or otheractive pharmaceutical ingredients (APIs) within the compositions. Itwill be understood that any suitable combination of excipients may bepresent in the pharmaceutical compositions described herein.

Excipients for use in topical formulations are well-known in the art andexamples may be found in the Handbook of Pharmaceutical Excipients(Rowe, R. C. et al., APhA Publications; 5th ed., 2005). Classes ofexcipients may include waxes, emollients, thickening agents/viscosityincreasing agents, humectants, pH modifiers, water repelling agents,anti-foaming agents, surfactants, solubilizers, wetting agents,penetration enhancers, antioxidants, and solvents. The excipients mayalso be present in the topical composition at any suitableconcentration. In some embodiments, the topical composition includesexcipients at a concentration in a range from 70 to 99.99 weightpercent.

Any suitable viscosity increasing agent may be used, and combinations ofviscosity increasing agents may also be used. In some embodiments of theinvention, the polymeric portion of the viscosity increasing agent mayact as a visco-elastic substance and may retain the gel at the site ofapplication, along with the APIs dispersed therein. Examples ofviscosity increasing agents include co-polymers ofcarboxymethylcellulose and acrylic acid, N-vinylpyrrolidone,polyalkylene glycols (e.g., poly(ethylene glycol)), polyalkylene oxides(e.g., polyethylene oxide), polyvinyl alcohols, polyvinylpyrrolidone,polysiloxanes, poly(vinyl acetates), cellulose, derivatized celluloses,alginates, copolymers thereof and blends thereof. A specific example ofa viscosity agent is hydroxypropylcellulose, such as Klucel®hydroxypropylcellulose (e.g., Klucel® MF Pharm grade).

Any suitable solvent or combinations of solvents may be used in thetopical compositions. Examples of solvents include acetone, methylalcohol, ethanol, isopropanol, butyl alcohol, ethyl acetate, dimethylisosorbide, propylene glycol, glycerol, ethylene glycol, polyethyleneglycol, diethylene glycol monoethyl ether or mixtures thereof. Inparticular examples, the solvent includes ethanol. In some embodiments,the solvent includes isopropyl alcohol. The skilled artisan willappreciate that the solvents may also be considered excipients,particularly at lower concentrations.

Any suitable humectant or combination of humectants may be used.Examples include glycols, such as diethylene glycol monoethyl ether;glycerols; sugar polyols, such as sorbitol, xylitol and maltitol;polyols such as polydextroses; quillaia, urea, and blends thereof. Inparticular examples, the humectant includes an alkylene glycol, such ashexylene glycol.

In some embodiments, the anhydrous topical compositions include at leastone water repelling agent, also referred to as a water repellant.Examples of water repelling agents include silicones, such ascyclomethicone, dimethicone, simethicone, C26-28 alkyl dimethicone,C26-28 alkyl methicone, polyphenylsisquioxane, trimethylsiloxysilicateand crosspolymers of cyclopentasiloxane anddimethicone/vinyltrimethylsiloxysilicate, and blends thereof. The waterrepelling agent may be particularly useful in embodiments where thetopical vehicle is used with a water-reactive API, such as a nitricoxide-releasing API whereby the nitric oxide is released in the presenceof water (e.g., a diazeniumdialate). In other cases, such as when theAPI is not water sensitive, a water repelling agent may not be included.

In some embodiments of the invention, the anhydrous topical compositionsmay have a viscosity increasing agent concentration in a range from 0.5to 30% by weight, a solvent concentration in a range from 50 to 90% byweight, and a humectant concentration in a range from 2 to 20% byweight. In some embodiments, the anhydrous topical compositions may alsoinclude a water repelling agent at a concentration in a range from 0.5%to 15% by weight. In some embodiments, the topical anhydrouscompositions further include at least one API, for example, at aconcentration in a range from 0.01 to 30% by weight, or any rangetherein.

In particular embodiments, the solvent is ethyl alcohol, 200 proof,anhydrous, the humectant is hexylene glycol available from NexioSolutions; the water repelling agent is ST-cyclomethicone-5 NF availablefrom Dow Corning Corp.; and the viscosity increasing agent is ahydroxypropylcellulose (HPC), such as Klucel® MF Pharm available fromAshland Aqualon.

In specific embodiments, the vehicle has the formulation of about 83.5%ethyl alcohol as a solvent, about 10% hexylene glycol as a humectant,about 2.5% ST-cyclomethicon-5 as a water repelling agent, about 2% HPCas a viscosity increasing agent, leaving about 2% available forinclusion of an API. In other embodiments the API is a nitric oxidedonor. In other embodiments, the API is Nitricil™, a NO-releasingco-condensed silica, from Novan, Inc. In some embodiments, the totalnitric oxide loading in the topical gel may be in a range from 0.01 to5.0 wt/wt %. In some embodiments, the total nitric oxide loading in thetopical gel is in a range from 0.24 to 0.36 wt/wt %.

In particular embodiments, the solvent is isopropyl alcohol, USP, thehumectant is hexylene glycol available from Nexio Solutions; the waterrepelling agent is ST-cyclomethicone-5 NF available from Dow CorningCorp.; and the viscosity increasing agent is a hydroxypropylcellulose(HPC), such as Klucel® MF Pharm available from Ashland Aqualon.

In particular embodiments, the vehicle has the formulation of 86.5% lessthe percentage of API of isopropyl alcohol as a solvent, 10% hexyleneglycol as a humectant, 2.5% ST-cyclomethicon-5 as a water repellingagent, and 1% HPC as a viscosity increasing agent. Decreasing thequantity of viscosity increasing agent in the formulation may allow forhigher loadings of API while maintaining acceptable organolepticproperties, such as not feeling gritty. The quantity of API may beanywhere from 0.01% to 50% or any range therein, such as, but notlimited to, from 0.1% to 30% or from 2% to 20%. In certain embodimentsthe API is a nitric oxide donor. In other embodiments, the API isNitricil™, a NO-releasing co-condensed silica, from Novan, Inc. In someembodiments, the total nitric oxide loading in the topical gel may be ina range from 0.01 to 30 wt/wt % or any range therein, such as, but notlimited to, 0.1 to 15 wt/wt %. In some embodiments, the total nitricoxide loading in the topical gel is in a range from 0.1 to 5 wt/wt % orany range therein. The total nitric oxide loading in a topical gel maybe measured by methods known to those of skill in the art, including,but not limited to, using a nitric oxide analyzer, such as a Sievers280i Nitric Oxide Analyzer.

For applications where the API is water reactive, such as a nitricoxide-releasing particle that releases nitric oxide in the presence ofwater, the inclusion of the water repelling agent may increase the APIstorage stability of the topical gel. In some embodiments, uniformlydispersing a water repellant with the API in a ratio of API to waterrepellant of from 0.01:1 to 30:1 and, in particular embodiments, 2 to 1,4 to 1, 6 to 1, 8 to 1, 12 to 1 or 20 to 1, may, similarly, allow closeassociation of the water repellant with the API and, thereby, improvestability of the topical gel.

For applications where the topical composition must absorb water,dissociate when in contact with an aqueous environment (e.g. on thesurface of the skin), or utilize water as a mechanism for initiatingdrug release/delivery, the inclusion of a humectant may be used. Forexample, where the API is a diazeniumdiolated co-condensed silicaparticle, the humectant may be provided in a ratio of humectant to APIof from 1:3 to 20:1 by weight to the API and, in some embodiments at aratio from 5 to 1, 5 to 3, 1 to 1, 1 to 1.2 or 1 to 2 to the API.Uniformly dispersing the humectant with the API may allow for the closeproximity of the humectant with the API in the final topical gel and,thereby, provide hydrophilic channels or access to moisture uptake in ananhydrous topical gel.

In compositions with water sensitive APIs (i.e., diazeniumdiolate),cellulose polymers may be useful as viscosity increasing agents becausethe crosslinking of the gel presents both a physical barrier to moisturediffusion and a hydrogen bonding network of water to protect it fromgetting to the water reactive functional groups. However, the cellulosepolymer may not inhibit the release of nitric oxide in a moisture richenvironment, such as on the skin of a patient. The degree of stabilityenhancement may be based on the weight percent of the cellulose polymer.Additional factors affecting nitric oxide stability may include themolecular weight of the cellulose. In some embodiments, the topicalcompositions may be considered pharmaceutically acceptable. Apharmaceutically acceptable composition, as defined herein, refers to acomposition that is suitable for application to a subject, such as ahuman, without undue side effects such as toxicity or irritation to theskin. Undue side effects are those that render the compositionunsuitable for application to a subject because the harm from the sideeffects outweighs the benefits of the composition.

In embodiments wherein the API includes a NO-releasing compound, theNO-releasing compounds may be present in pharmaceutically acceptablecompositions according to embodiments of the invention at any suitableconcentration, but in some embodiments, the NO-releasing compounds arepresent in the compositions at a concentration sufficient to elicit animprovement in, eliminate or prevent acne. In some embodiments, theconcentration of NO-releasing compounds ranges from 0.1% to 30% w/w inthe composition. In particular embodiments, the concentration of theNO-releasing compounds is less than 20% w/w in the composition or anyrange therein. In still further embodiments, the concentration of theNO-releasing compounds is less than 10% w/w in the composition or anyrange therein. In particular embodiments, the concentration of theNO-releasing compounds is 1%, 2%, 4%, or 8% w/w in the composition.

While pharmaceutically acceptable compositions that may be used incertain embodiments of the present inventive concept, in particularembodiments, a topical gel is provided. Topical gels according to thepresent inventive concept may be non-irritating to the skin, may providenoticeable results in a relatively short period of time, may becosmetically elegant and/or may not bleach or otherwise stain clothingand other items that contact the product.

Additionally, to avoid a gritty feeling of the topical gel, the maximumparticle size for a the API may be less than 100 μm, or any rangetherein, and, in some embodiments, less than 20 μm, and in furtherembodiments, less than 10 μm.

The topical vehicle and API may be readily spread by a patient and,therefore, may have a viscosity of from 250 cP to 50000 cP or any rangetherein, such as, but not limited to, 500 cP to 10000 cP or 1000 cP to8000 cP. In some embodiments, the viscosity of the topical gel is about7000 cP. The viscosity of the gel may be controlled by the amount of theviscosity increasing agent added to the gel. Too much of the agent andthe gel will become too thick and difficult to spread.

The topical gel may also evaporate relatively quickly to avoid a feelingof greasiness and, in some cases, to provide a cooling sensation. Thus,the topical gel may have a relatively high level of volatiles. In someembodiments, the percent of volatiles by weight is greater than 55% orany range therein. In particular embodiments, the percent of volatilesby weight is about 55%, 66%, 74%, 80%, or 86%.

Additionally, the topical gel may have a “leave-on” layer or secondaryformulation after evaporation of the volatiles that is spreadsubstantially uniformly when the topical gel is spread on the skin. Thepercent non-volatiles may be from 2 to 45% by weight to provide asuitable leave-on layer. In some embodiments, the percent non-volatilesis about 14%, 17%, 23%, 31% or 41% by weight. In some embodiments, theleave-on layer comprises nitric oxide-releasing particles,hydroxypropylcellulose, and hexylene glycol.

A topical gel of the present invention may have a pH from 5 to 12, orany range therein, such as, but not limited to, from 7 to 12, 8 to 11,or 10 to 11. In certain embodiments, a topical gel may have a pH ofabout 5, 6, 7, 8, 9, 10, 11, or 12. In some embodiments, such as, butnot limited to, embodiments comprising a water reactive API, the pH of atopical gel may change upon use (e.g., contact with the skin of asubject) and/or upon contact with water.

As described herein, selection of the ingredients may affect thestability and, therefore, the release of the nitric oxide from the API.Accordingly, the selected ingredients and their proportions may beselected to provide a desired stability and release profile for a givennitric oxide-releasing particle. Furthermore, as described below, theprocessing of the API and vehicle may also impact stability and releasekinetics. For diazeniumdiolate NO-releasing APIs, considerations indetermining the impact of the vehicle constituents and the processinginclude rate of release of nitric oxide desired, including the releaseprofile desired, the type of ailment that the composition aims to treat,other APIs included in the composition and the stability desired.

Active Pharmaceutical Ingredients

Any suitable active pharmaceutical ingredient (API) or combinations ofAPIs may be included in the compositions according to embodiments of theinvention. Examples of APIs include antimicrobial agents, anti-acneagents, anti-inflammatory agents, analgesic agents, anesthetic agents,antihistamine agents, antiseptic agents, immunosuppressants,antihemorrhagic agents, vasodilators, wound healing agents, anti-biofllmagents and mixtures thereof.

Examples of antimicrobial agents include penicillins and related drugs,carbapenems, cephalosporins and related drugs, erythromycin,aminoglycosides, bacitracin, gramicidin, mupirocin, chloramphenicol,thiamphenicol, fusidate sodium, lincomycin, clindamycin, macrolides,novobiocin, polymyxins, rifamycins, spectinomysin, tetracyclines,vanomycin, teicoplanin, streptogramins, anti-folate agents includingsulfonamides, trimethoprim and its combinations and pyrimethamine,synthetic antibacterials including nitrofurans, methenamine mandelateand methenamine hippurate, nitroimidazoles, quinolones,fluoroquinolones, isoniazid, ethambutol, pyrazinamide,para-aminosalicylic acid (PAS), cycloserine, capreomycin, ethionamide,prothionamide, thiacetazone, viomycin, eveminomycin, glycopeptide,glyclyclycline, ketolides, oxazolidinone; imipenen, amikacin,netilmicin, fosfomycin, gentamycin, ceftriaxone, Ziracin, Linezolid,Synercid, Aztreonam, and Metronidazole, Epiroprim, Sanfetrinem sodium,Biapenem, Dynemicin, Cefluprenam, Cefoselis, Sanfetrinem celexetil,Cefpirome, Mersacidin, Rifalazil, Kosan, Lenapenem, Veneprim, Sulopenem,ritipenam acoxyl, Cyclothialidine, micacocidin A, carumonam, Cefozopranand Cefetamet pivoxil.

Examples of topical anti-acne agents include adapalene, azelaic acid,benzoyl peroxide, clindamycin and clindamycin phosphate, doxycycline,erythromycin, keratolytics such as salicylic acid and retinoic acid(Retin-A″), norgestimate, organic peroxides, retinoids such asisotretinoin and tretinoin, sulfacetamide sodium, and tazarotene.Particular anti-acne agents include adapalene, azelaic acid, benzoylperoxide, clindamycin (e.g., clindamycin phosphate), doxycycline (e.g.,doxycycline monohydrate), erythromycin, isotretinoin, norgestimate,sulfacetamide sodium, tazarotene, etretinate and acetretin.

Examples of antihistamine agents include diphenhydramine hydrochloride,diphenhydramine salicylate, diphenhydramine, chlorpheniraminehydrochloride, chlorpheniramine maleate isothipendyl hydrochloride,tripelennamine hydrochloride, promethazine hydrochloride, methdilazinehydrochloride, and the like. Examples of local anesthetic agents includedibucaine hydrochloride, dibucaine, lidocaine hydrochloride, lidocaine,benzocaine, p-buthylaminobenzoic acid 2-(die-ethylamino)ethyl esterhydrochloride, procaine hydrochloride, tetracaine, tetracainehydrochloride, chloroprocaine hydrochloride, oxyprocaine hydrochloride,mepivacaine, cocaine hydrochloride, piperocaine hydrochloride, dyclonineand dyclonine hydrochloride.

Examples of antiseptic agents include alcohols, quaternary ammoniumcompounds, boric acid, chlorhexidine and chlorhexidine derivatives,iodine, phenols, terpenes, bactericides, disinfectants includingthimerosal, phenol, thymol, benzalkonium chloride, benzethoniumchloride, chlorhexidine, povidone iode, cetylpyridinium chloride,eugenol and trimethylammonium bromide.

Examples of anti-inflammatory agents include nonsteroidalanti-inflammatory agents (NSAIDs); propionic acid derivatives such asibuprofen and naproxen; acetic acid derivatives such as indomethacin;enolic acid derivatives such as meloxicam, acetaminophen; methylsalicylate; monoglycol salicylate; aspirin; mefenamic acid; flufenamicacid; indomethacin; diclofenac; alclofenac; diclofenac sodium;ibuprofen; ketoprofen; naproxen; pranoprofen; fenoprofen; sulindac;fenclofenac; clidanac; flurbiprofen; fentiazac; bufexamac; piroxicam;phenylbutazone; oxyphenbutazone; clofezone; pentazocine; mepirizole;tiaramide hydrochloride; steroids such as clobetasol propionate,bethamethasone dipropionate, halbetasol proprionate, diflorasonediacetate, fluocinonide, halcinonide, amcinonide, desoximetasone,triamcinolone acetonide, mometasone furoate, fluticasone proprionate,betamethasone diproprionate, triamcinolone acetonide, fluticasonepropionate, desonide, fluocinolone acetonide, hydrocortisone vlaerate,prednicarbate, triamcinolone acetonide, fluocinolone acetonide,hydrocortisone and others known in the art, predonisolone,dexamethasone, fluocinolone acetonide, hydrocortisone acetate,predonisolone acetate, methylpredonisolone, dexamethasone acetate,betamethasone, betamethasone valerate, flumetasone, fluorometholone,beclomethasone diproprionate, fluocinonide, topical corticosteroids, andmay be one of the lower potency corticosteroids such as hydrocortisone,hydrocortisone-21-monoesters (e.g., hydrocortisone-21-acetate,hydrocortisone-21-butyrate, hydrocortisone-21-propionate,hydrocortisone-21-valerate, etc.), hydrocortisone-17,21-diesters (e.g.,hydrocortisone-17,21-diacetate, hydrocortisone-17-acetate-21-butyrate,hydrocortisone-17,21-dibutyrate, etc.), alclometasone, dexamethasone,flumethasone, prednisolone, or methylprednisolone, or may be a higherpotency corticosteroid such as clobetasol propionate, betamethasonebenzoate, betamethasone dipropionate, diflorasone diacetate,fluocinonide, mometasone furoate, triamcinolone acetonide.

Examples of analgesic agents include alfentanil, benzocaine,buprenorphine, butorphanol, butamben, capsaicin, clonidine, codeine,dibucaine, enkephalin, fentanyl, hydrocodone, hydromorphone,indomethacin, lidocaine, levorphanol, meperidine, methadone, morphine,nicomorphine, opium, oxybuprocaine, oxycodone, oxymorphone, pentazocine,pramoxine, proparacaine, propoxyphene, proxymetacaine, sufentanil,tetracaine and tramadol.

Examples of anesthetic agents include alcohols such as phenol; benzylbenzoate; calamine; chloroxylenol; dyclonine; ketamine; menthol;pramoxine; resorcinol; troclosan; procaine drugs such as benzocaine,bupivacaine, chloroprocaine; cinchocaine; cocaine; dexivacaine;diamocaine; dibucaine; etidocaine; hexylcaine; levobupivacaine;lidocaine; mepivacaine; oxethazaine; prilocaine; procaine; proparacaine;propoxycaine; pyrrocaine; risocaine; rodocaine; ropivacaine; tetracaine;and derivatives, such as pharmaceutically acceptable salts and estersincluding bupivacaine HCl, chloroprocaine HCl, diamocaine cyclamate,dibucaine HCl, dyclonine HCl, etidocaine HCl, levobupivacaine HCl,lidocaine HCl, mepivacaine HCl, pramoxine HCl, prilocaine HCl, procaineHCl, proparacaine HCl, propoxycaine HCl, ropivacaine HCl, and tetracaineHCl.

Examples of antihemorrhagic agents include thrombin, phytonadione,protamine sulfate, aminocaproic acid, tranexamic acid, carbazochrome,carbaxochrome sodium sulfanate, rutin and hesperidin.

In some embodiments of the invention, the active pharmaceuticalingredient (API) includes a compound that releases nitric oxide (NO).Any suitable NO-releasing compound may be used. In some embodiments, theNO-releasing compound includes a small molecule compound that includesan NO donor group. Small molecule compounds are defined herein ascompounds having a molecular weight of less than 500 daltons, andinclude organic and/or inorganic small molecules. In some embodiments,the NO-releasing compound includes a macromolecule that includes an NOdonor group. A macromolecule is defined herein as any compound that hasa molecular weight of 500 daltons or greater. Any suitable macromoleculemay be used, including crosslinked or non-crosslinked polymers,dendrimers, metallic compounds, organometallic compounds,inorganic-based compounds, and other macromolecular scaffolds. In someembodiments, the macromolecule has a nominal diameter ranging from 0.1nm-100 μm and may comprise the aggregation of two or moremacromolecules, whereby the macromolecular structure is further modifiedwith a NO donor group.

In some embodiments of the invention, the NO donor of the NO-releasingcompound releases nitric oxide upon exposure to an external condition,such as light, heat, water, acid, base, and the like. For example, insome embodiments, the NO-releasing compound includes a diazeniumdiolatefunctional group as an NO donor. The diazeniumdiolate functional groupmay produce nitric oxide under certain conditions, such as upon exposureto water. As another example, in some embodiments, the NO-releasingcompound includes a nitrosothiol functional group as the NO donor. TheNO donor may produce nitric oxide under certain conditions, such as uponexposure to light. Examples of other NO donor groups includenitrosamine, hydroxyl nitrosamine, hydroxyl amine and hydroxyurea. Anysuitable combination of NO donors and/or NO-releasing compounds may alsobe used in the methods described herein. Additionally, the NO donor maybe incorporated into or onto the small molecule or macromolecule throughcovalent and/or non-covalent interactions.

In some embodiments of the invention, the NO-releasing compounds may bein the form of NO-releasing particles, such as those described in U.S.Publication No. 2009/0214618, the disclosure of which is incorporated byreference herein in its entirety. Such particles may be prepared bymethods described therein.

The NO-releasing compound may release nitric oxide by any suitablemechanism, including via reaction with water and/or thermal degradation.Examples of NO-releasing functional groups that may be included in theNO-releasing compound include, but are not limited to, diazeniumdiolate,nitrosamine, hydroxyl nitrosamine, nitrosothiol, hydroxyl amine,hydroxyurea, and metal nitrosyl complexes. Other NO-releasing functionalgroups that are capable of releasing nitric oxide in a therapeuticmanner, such as acidified nitrite, may also be utilized.

The NO-releasing compound may be a small molecule compound, an oligomerand/or a polymer and may be in any suitable physical form, such as, butnot limited to, a particle, coating, film, liquid, solution and thelike. In some embodiments, the nitric oxide-releasing compound comprisesdiazeniumdiolate-functionalized polysiloxane macromolecules as describedabove. Other non-limiting examples of NO-releasing compounds includeNO-releasing zeolites as described in United States Patent PublicationNos. 2006/0269620 or 2010/0331968; NO-releasing metal organic frameworks(MOFs) as described in United States Patent Application Publication Nos.2010/0239512 or 2011/0052650; NO-releasing multi-donor compounds asdescribed in U.S. Provisional Patent Application Ser. No. 61/526,918entitled “Tunable Nitric Oxide-Releasing Macromolecules Having MultipleNitric Oxide Donor Structures”; NO-releasing dendrimers or metalstructures as described in U.S. Publication No. 2009/0214618; nitricoxide releasing coatings as described in U.S. Publication No.2011/0086234; and compounds as described in U.S. Publication No.2010/0098733. The disclosures of each of the references in thisparagraph are incorporated herein by reference in their entirety.Additionally, NO-releasing macromolecules may be fabricated as describedin International Application No. PCT/US2012/022048 entitled “TemperatureControlled Sol-Gel Co-Condensation” filed Jan. 20, 2012, the disclosureof which is incorporated herein by reference in its entirety.

As an example, in some embodiments of the invention, the NO-releasingparticles include NO-loaded precipitated silica. The NO-loadedprecipitated silica may be formed from nitric oxide donor modifiedsilane monomers into a co-condensed siloxane network. In one embodimentof the invention, the nitric oxide donor is an N-diazeniumdiolate.

In some embodiments, the nitric oxide donor may be formed from anaminoalkoxysilane by a pre-charging method, and the co-condensedsiloxane network may be synthesized from the condensation of a silanemixture that includes an alkoxysilane and the aminoalkoxysilane to forma nitric oxide donor modified co-condensed siloxane network. As usedherein, the “pre-charging method” means that aminoalkoxysilane is“pretreated” or “precharged” with nitric oxide prior to theco-condensation with alkoxysilane. In some embodiments, the prechargingnitric oxide may be accomplished by chemical methods. In anotherembodiment, the “pre-charging” method may be used to create co-condensedsiloxane networks and materials more densely functionalized withNO-donors.

The co-condensed siloxane network may be silica particles with a uniformsize, a collection of silica particles with a variety of size, amorphoussilica, a fumed silica, a nanocrystalline silica, ceramic silica,colloidal silica, a silica coating, a silica film, organically modifiedsilica, mesoporous silica, silica gel, bioactive glass, or any suitableform or state of silica.

In some embodiments, the alkoxysilane is a tetraalkoxysilane having theformula Si(OR)4, wherein R is an alkyl group. The R groups may be thesame or different. In some embodiments the tetraalkoxysilane is selectedas tetramethyl orthosilicate (TMOS) or tetraethyl orthosilicate (TEOS).In some embodiments, the aminoalkoxysilane has the formula:R″—(NH—R′)n-Si(OR)3, wherein R is alkyl, R′ is alkylene, branchedalkylene, or aralkylene, n is 1 or 2, and R″ is selected from the groupconsisting of alkyl, cycloalkyl, aryl, and alkylamine.

In some embodiments, the aminoalkoxysilane may be selected fromN-(6-aminohexyl)aminopropyltrimethoxysilane (AHAP3);N-(2-aminoethyl)-3-aminopropyltrimethoxysilane (AEAP3);(3-trimethoxysilylpropyl)di-ethylenetriamine (DET3);(aminoethylaminomethyl)phenethyltrimethoxysilane (AEMP3);[3-(methylamino)propyl]trimethoxysilane (MAP3);N-butylamino-propyltrimethoxysilane(n-BAP3);t-butylamino-propyltrimethoxysilane(t-BAP3);N-ethylaminoisobutyltrimethoxysilane(EAiB3); N-phenylamino-propyltrimethoxysilane (PAP3); andN-cyclohexylaminopropyltrimethoxysilane (cHAP3).

In some embodiments, the aminoalkoxysilane has the formula:NH[R′—Si(OR)3]2, wherein R is alkyl and R′ is alkylene. In someembodiments, the aminoalkoxysilane may be selected frombis(3-triethoxysilylpropyl)amine, bis-[3-(trimethoxysilyl)propyl]amineand bis-[(3-trimethoxysilyl)propyl]ethylenediamine.

In some embodiments, as described herein above, the aminoalkoxysilane isprecharged for NO-release and the amino group is substituted by adiazeniumdiolate. Therefore, in some embodiments, the aminoalkoxysilanehas the formula: R″—N(NONO—X+)—R′—Si(OR)3, wherein R is alkyl, R′ isalkylene or aralkylene, R″ is alkyl or alkylamine, and X+ is a cationselected from the group consisting of Na+, K+ and Li+.

The composition of the siloxane network, (e.g., amount or the chemicalcomposition of the aminoalkoxysilane) and the nitric oxide chargingconditions (e.g., the solvent and base) may be varied to optimize theamount and duration of nitric oxide release. Thus, in some embodiments,the composition of the silica particles may be modified to regulate thehalf-life of NO release from silica particles.

In another embodiment, the amino group of aminoalkoxysilane issubstituted with a diazeniumdiolate, and the aminoalkoxysilane having aformula of R″—N(NONO—X+)—R′—Si(OR)3, wherein: R is alkyl, R′ is alkyleneor aralkylene, R″ is alkyl or alkylamine, and X+ is a cation selectedfrom the group consisting of Na+ and K+.

In some embodiments of the invention, the particle size of theNO-releasing particles is in a range from 20 nm to 10 μm. The particlesize may be tailored to minimize or prevent toxicity and penetrationthrough the epidermis (or compromised dermis) and into the bloodvessels. In particular embodiments, the particle size is distributedaround a mean particle size of less than 10 μm, or any range therein, toallow the particle to enter a follicle. In further embodiments, theparticle size is distributed around a mean particle size of less than 8μm, or any range therein. In other embodiments, the particle size isdistributed around a mean particle size of greater than 10 μm, or anyrange therein, to prevent the particle from entering the follicle.

In still further embodiments, a mixture of particles with mean particlesizes distributed around two or more mean particle sizes may beprovided. For example, a mixture of particles having a particle sizedistributed around a mean particle size of less than 10 μm to allow theparticle to enter a follicle may be mixed with particles having aparticle size distributed around a mean particle size of greater than 10μm to prevent the particle from entering the follicle. The particles mayhave the same nitric oxide release profiles or different nitric oxiderelease profiles. For example, the smaller particles may have a releaseprofile tailored to enhance their ability to moderate sebum productionand/or abnormal keratinization and the larger particles may have arelease profile tailored to enhance their ability to kill bacteria,promote wound healing, reduce scarring or other desirable therapeuticeffect provided by nitric oxide. Other combinations and multiplecombinations could also be provided.

Synthetic Methods and Apparatus

The compositions described herein may be made by any suitable method.However, in some embodiments, the compositions may be manufactured by amethod described herein and in U.S. Provisional Patent Application Ser.Nos. 61/504,626 and 61/610,179, both entitled “Methods of ManufacturingTopical Compositions and Apparatus For Same,” filed Jul. 5, 2011 andMar. 13, 2012, respectively, which are hereby incorporated by referenceherein in their entirety.

In some embodiments, such methods include homogenizing a first excipientcomposition that includes a viscosity increasing agent and at least onesolvent to form a first premix composition; separately homogenizing atleast one active pharmaceutical ingredient (API) and a second excipientcomposition to form a second premix composition; and combining the firstpremix composition and the second premix composition to form theanhydrous topical composition. The pre-mixing of constituents mayincrease the stability of the API and provide uniformity of thedispersion of the constituents throughout the final topical composition.In some embodiments, the first excipient composition includes at leastone viscosity agent, at least one solvent and at least one humectant,while the second excipient composition includes at least one waterrepellant and at least one humectant. In some cases, the humectant maybe divided between the first excipient composition and the secondexcipient composition so that the humectant may aid in swelling of theviscosity increasing agent.

Any suitable homogenization mechanism may be used. Examples ofhomogenization devices include mechanical overhead agitation such aspropeller, anchor, pitch blade, rotor-stator, rotating blades,ultrasonic devices, in-line and high pressure homogenizers. Any of thesemethods may be used, and multiple methods may be used in combination insome embodiments. Homogenization of the premix compositions may providea final topical composition that has desirable API stability and blendhomogeneity. In some embodiments of the present invention, an in-linehomogenizer may be used. Homogenization may be carried out continuouslythroughout out the mixing of a composition and/or portion of acomposition (e.g., a first and/or second excipient composition). In someembodiments, homogenization may be carried out intermittently throughoutout the mixing of a composition and/or portion of a composition (e.g., afirst and/or second excipient composition). In certain embodiments,homogenization may be carried out intermittently in durations of 15, 14,13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 minutes, or any rangetherein, every 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1minutes, or any range therein.

In particular embodiments of the present invention, a homogenizationmethod and/or device may be used that maintains the activepharmaceutical ingredient (e.g., a water reactive active pharmaceuticalingredient) below a temperature at which the active pharmaceuticalingredient could degrade. An active pharmaceutical ingredient maydegrade at a particular temperature if maintained at that temperaturefor a specific duration of time. Accordingly, in some embodiments of thepresent invention, the duration of time an active pharmaceuticalingredient is maintained at a particular temperature, is below the timeperiod at which the active ingredient could degrade at that temperature.In certain embodiments of the present invention, for the entirehomogenization process, the active pharmaceutical ingredient is kept ata temperature that does not exceed the temperature at which the activepharmaceutical ingredient could degrade.

In some embodiments, homogenization is performed at a temperature in arange from −15° C. to 30° C., or any range therein, such as, but notlimited to, −5° C. to 25° C., 0° C. to 10° C., 10° C. to 20° C., or 15°C. to 25° C. In some embodiments, the homogenization is performed atroom temperature. In other embodiments, the homogenization is performedbelow room temperature.

In some embodiments, homogenization is performed in a dry, inertatmosphere, such that water and oxygen are substantially absent from thehomogenization vessel.

In some embodiments, homogenizing the first excipient composition thatincludes the at least one viscosity increasing agent and the at leastone solvent includes adding the viscosity increasing agent to thesolvent at a relatively rapid rate, such as at a rate in a range of 0.5to 50 g of viscosity increasing agent per min per liter solvent. In oneembodiment, the rate is 15 g of viscosity increasing agent per min perliter solvent. In some embodiments where a humectant is added to thefirst excipient composition, a portion of the humectant may behomogenized with the at least one solvent, and then the at least oneviscosity increasing agent may be added to the mixture and the firstexcipient composition may be homogenized. In other embodiments, aportion of the humectant, at least one solvent, and at least oneviscosity increasing agent are added and homogenized together at thesame time. One or more portions of the humectant, such as 1, 2, 3, ormore, may be added to the first excipient composition at differenttimes. Accordingly, when a humectant is added to the first excipientcomposition, the humectant may be portioned into 2, 3, 4, or more equaland/or unequal portions, and one or more portions of the humectant maybe added to the first excipient composition. For example, a humectantmay be divided into three equal and/or unequal portions, and two of thethree portions may be added to the first excipient composition atdifferent times with the remaining portion added to the second excipientcomposition.

The rapid addition of a viscosity increasing agent such as hydroxypropylcellulose to the solvent is contrary to the typical practice in theindustry that would suggest slow addition of a viscosity increasingagent such as hydroxypropyl cellulose to the solvent to avoid clumping.In contrast, the present inventors have found that by using thehomogenization apparatus described herein as a premixing vessel, theviscosity increasing agent may be rapidly added. In fact, in some cases,rapid addition may result in a more homogeneous mixture. In someembodiments, the homogenization of the first excipient composition andthe at least one solvent may be performed at a rate of 2,500 to 10,000RPM, for example, with an IKA T-50 UltraTurax rotor stator device orequivalent mixing conditions utilizing alternative homogenizationdevices.

In some embodiments, homogenizing the second excipient composition isperformed at a rate of 2500 to 10,000 rpm. In a particular embodimentthe rate is 7,500 RPM utilizing the rotor stator configuration describedabove. Homogenization may be achieved by using an apparatus describedherein. In some embodiments of the invention, the homogenizationapparatus includes a vessel and a rotor-stator therein. In someembodiments, the ratio of the height of the reactive portion of thevessel to the width of the vessel is in a range of 1:2 to 1:1. In someembodiments, the widest portion of the rotor-stator has a diameter thatis 5 to 50% of the width of the vessel. The vessel may be temperaturecontrolled and may include an inlet and outlet for an inert gas. Thevessel may be made of any suitable material, including, for example,glass, stainless steel, polytetrafluoroethylene, and other non-reactivematerials. Particular embodiments of the homogenization apparatus willbe described in detail below.

Once the two premix compositions are prepared, they may be combined andmixed. In some embodiments, the premix compositions are mixed at a ratein a range of 10 to 50 RPM. In addition to mixing the two premixcompositions, the combined composition may be homogenized, for example,under high shear using IKA Labor Pilot 2000/4 in-line homogenizeroperating at a rate in a range of 2,000 to 24,000 RPM, allowing materialto be recirculated at a rate of up to 4 gallons per minute, orequivalent conditions with alternative homogenization devices. Such ahomogenization process may smooth, deagglomerate, reduce particle size,increase blend uniformity of excipient and active pharmaceuticalingredients, improve viscosity uniformity, reduce segregation ofsuspended particulates, and/or improve API stability.

Process parameters for an exemplary homogenization process are providedin Table 1.

TABLE 1 Exemplary process parameters Process Parameter Setting/RangePre-mixing vessel temperature 18° C. to 22° C. Main vessel temperature18° C. to 22° C. Inert gas flow rate about 0.2 liters per minute Rate ofaddition for hydroxypropyl  1 minute or less cellulose Mixer/agitatorspeed setting for 75 Hz to 90 Hz the main vessel Mixing time for thefirst excipient 30 minutes or more composition In-line homogenizationtime for the homogenize in about 5 minute first excipient compositiondurations every 5 to 10 minutes throughout the mixing time for the firstexcipient composition Rotor-stator homogenization time for  2 minutes ormore the second excipient composition Mixing time for the composition 30minutes or more In-line homogenization time for the homogenize in about2 minute composition durations every 5 to 10 minutes throughout themixing time for the composition

Exemplary Methods and Apparatus for Manufacturing Topical Compositions

FIG. 1 is a detailed drawing of a system 100 for the manufacture offormulations according to some embodiments of the present invention. Asseen in FIG. 1, the system 100 includes a pre-mixing vessel 105 that isprovided by a rotor-stator homogenizer 110 in a single jacketed vessel120 that is connected to a circulator 130 for temperature control. Thejacketed vessel 120 may be operated under an blanket of inert gas, suchas argon, with a rotameter 230 used to control the rate of inert gasaddition by controlling the flow from one of the two outputs of athree-way valve. A vent line 150 is also present to exhaust excess inertgas. Raw materials are introduced from holding vessels 50 to thepre-mixing vessel 105.

The pre-mixing vessel 105 is used for two primary purposes in the system100: (1) to provide a forced wetting mechanism for excipients such as,but not limited to, viscosity increasing agents such as hydroxypropylcellulose (HPC) in a suspension consisting of solvents and excipients toproduce a gel, ointment or cream; and (2) to disperse an activepharmaceutical ingredient, such as those described herein, in asuspension of the active pharmaceutical ingredient and solvents orexcipients. The pre-mixing vessel 105 has an oversize valve 125 toprevent blockages during transfer of suspensions to a mixing vessel(kettle) 160. For example, the mixing vessel 160 may be a 15 Liter topdriven mixing vessel available from Lee Industries. The transfer ofsuspensions between the pre-mixing vessel 105 and the mixing vessel 160is carried out by using a peristaltic pump 155 at a controlled rate. Theperistaltic pump may, for example, be a Masterflex LS available fromCole-Parmer, or similar. The transfer rate of excipients into the system100 for pre-mixing may also be important to compounding in the mixingvessel 160.

The mixing vessel 160 includes kettle 161, a motor 170 to drive anoverhead anchor agitator with paddles 180 and has an in-line homogenizer190 that is connected to the bottom ball valve 165 of the mixing vessel160. The in-line homogenizer 190 may, for example, be a Labor Pilot2000/4 available from IKA-Werk, or similar. The mixing vessel 160includes a jacket 200 that is connected to a circulator 210 to maintaintemperature control. The mixing vessel 160 also has a RTD sensor 166 tomonitor temperature in the kettle 161 and is recorded automaticallyusing a recorder 220, such as a Honeywell chart recorder. The contentsof the kettle 161 may also be under an inert gas blanket with arotometer 230 used to control the rate of inert gas addition. A ventline 167 is also present on the mixing vessel 160 to exhaust excessinert gas. The in-line homogenizer 190 is used for processintensification from an enhanced mixing perspective, particularly duringsemi-solids compounding. Semi-solids products may also be discharged viathe use of the in-line homogenizer 190 through the use of a three-wayvalve 240.

Additionally, as illustrated in FIG. 1, optional in-lineprocessing/analysis equipment 245 may be provided. For example, thein-line processing/analysis equipment 245 may include a viscometer tomeasure viscosity, a Ramen spectrometer, a pH meter, lighting devices topre-age or otherwise process a composition, a colorimeter to measure thecolor and/or uniformity of the gel or other such in-line analysisequipment.

The system 100 may allow for excipients to be dispersed uniformlywithout agglomeration and lumping. A homogeneous solution, suspension,gel, ointment, emulsion, and cream, or other pharmaceutical composition,may be manufactured using this device. Furthermore, the system 100 usingthe homogenizer allows for active pharmaceutical ingredients to bedispersed in a suspension to improve blend uniformity and thus preventagglomeration formation and potential segregation. The in-linehomogenizer 190 may be used to further aid the formation of a semi-solidformulation that is homogenous and uniform in terms of excipient andactive pharmaceutical ingredient distribution. In some embodiments, itmay be desirable to use an inert gas environment in both the pre-mixingvessel 105 and mixing vessel 160 is to prevent moisture from entering.As such, such an apparatus may be useful for making moisture-sensitive,or otherwise anhydrous, formulations.

Operation of the system 100 will now be described with reference to anexemplary manufacturing process for producing a topical composition.FIG. 2 is a flowchart illustration of steps/operations that may becarried out according to some embodiments of the present invention. Asseen in FIG. 2, the first excipient composition is premixed in thepre-mixing vessel 100 (block 1000) and transferred to the mixing vessel160 (block 1010), for example, by the peristaltic pump 155. The forcedwetting of the viscosity increasing agent with the homogenizer 110 ofthe pre-mixing vessel 105 may allow for the rapid combination ofmaterials which, otherwise, would take substantially longer to mix. Thecontents of the mixing vessel 160 are mixed and periodically homogenized(block 1020), for example, by recirculating the contents of the mixingvessel 160 through the in-line homogenizer 190.

In block 1030, the API and the second excipient composition are premixedand homogenized in the homogenizer 110. Premixing these constituents mayimprove the uniformity of them in the final gel as well as closelyassociating the API with excipients such as the humectant and the waterrepelling agent which, in the case of a water reactive API, may improvethe stability of the API in the final topical composition. The pre-mixedcontents of the homogenizer 110 are then transferred to the mixingvessel 160 via the peristaltic pump 155 (block 1040) and the contentsmixed in the mixing vessel 160 (block 1050). The premix homogenizer 110is then rinsed with the solvent and the contents transferred to themixing vessel 160 (block 1060). The contents of the mixing vessel arethen mixed until the desired uniformity is obtained (block 1070).

FIG. 3 is a more detailed flowchart of operations for a particularsystem 100 according to some embodiments of the present inventiveconcept. As seen in FIG. 3, operations begin by dispensing raw materialsfor use in the process (block 2100). In particular embodiments for theexemplary system of FIG. 1, the raw materials may be dispensed asmultiple portions of the same raw material. Such portions may be basedon the batch size and the dimensions of the particular equipmentutilized. Thus, what is described herein with reference to multiplesub-processes could be combined into fewer sub-process or moresub-processes depending on the particular equipment utilized and batchsize. In some embodiments, for the initial mixing operations, thedispensed raw materials are ethyl alcohol and/or isopropyl alcohol asthe solvent and Klucel® hydroxypropyl cellulose as the viscosityincreasing agent.

The temperature of the pre-mixing vessel 105 and the mixing vessel 160are set to 20° C.+/−2° C. via the jacket 120 and the recirculationsystem 130 and the jacket 200 and recirculating system 210 (block 2110).The pre-mixing vessel 105 and mixing vessel 160 are blanketed with argongas and the flow set through the valve 140 to provide about 0.2 litersof argon per minute (block 2120). The alcohol is added to the pre-mixingvessel 105 (block 2130) optionally with a portion of the humectanthexylene glycol (block 2135) and then the homogenization rate of thehomogenizer 110 is set and begun (block 2140). In the present example,the homogenizer rate is set to 7500 revolutions per minute to provide ahigh shear homogenization. After beginning homogenization of thealcohol, the hydroxypropyl cellulose is rapidly added to the pre-mixingvessel 105 (block 2150). In particular, the hydroxypropyl cellulose maybe added in less than 2 minutes or any range therein, in some cases, inless than 1 minute and, in some cases, in about 30 seconds. In someembodiments, at least one humectant, such as hexylene glycol, is addedwith the hydroxypropyl cellulose. For example, in some embodiments, aportion of the total humectant in the gel is added in the firstexcipient mixture while the remaining humectant is added with the secondexcipient composition. Any suitable amount of humectant may be added tothe first excipient composition, but in some embodiments, the amount ofhumectant included in the first composition is that which is needed toswell the viscosity increasing agent.

After the hydroxypropyl cellulose is added to the pre-mixing vessel 105,the homogenization rate is reduced and the mixture is pumped through theperistaltic pump 155 to the mixing vessel 160 (block 2160) at a rate ofat least 250 mL/min to avoid gelation during the transfer process. Inparticular, the mixture may be pumped at a rate of 500 mL/min. Themixing vessel 160 is set to mix the contents with a mixer speed of 10 to50 RPM, and in particular at about 40 rpm. In the present example, thehomogenizer rate is reduced to 4000 revolutions per minute. Aftertransferring the mixed ethyl alcohol and hydroxypropyl cellulose to themixing vessel 160, the homogenizer rate may be reset to the higher rateand the process begun again if more batches of material are required(block 2170). In the present example, the operations of block 2100through 2170 are repeated three times. The pre-mixing vessel 105 is thenrinsed with alcohol for 5 minutes and the contents transferred to themixing vessel 160 through the peristaltic pump 155 at a flow rate of 500mL/min (block 2175). Excipients and solvents are dosed either manuallyor through automated methods.

When the pre-mixing vessel 105 has been rinsed and transferred to themixing vessel 106, the mixing vessel 160 mix rate is increased to about45 RPM and the contents mixed for about 45 minutes (block 2180). Every10 to 20 minutes or any range therein (block 2200), the contents of themixing vessel 160 are recirculated through the valve 165, the in-linehomogenizer 190, the valve 240 and optionally, the in-line analysisequipment 245 (block 2190) for about 3 minutes but not so long as tocause an undesirable temperature increase resulting from the high shearhomogenization.

In block 2210, the pre-mixing of the API begins by adding the humectanthexylene glycol (in which in some embodiments, the remaining portion ofthe humectant is added—for example, one half of the total humectantconcentration may optionally have been included in the first excipientcomposition (block 2135)) and ST-cyclomethicone-5 to the pre-mixingvessel 105. The homogenization rate of the homogenizer 110 is set andbegun (block 2220). In the present example, the homogenizer rate is setto 4000 revolutions per minute. The materials are homogenized for about5 minutes and the API (in this example, Nitricil™, a nitricoxide-releasing co-condensed silica) is added to the pre-mixing vessel105 (block 2230). The homogenizer rate is reduced to 2500 revolutionsper minute and the contents of the pre-mixing vessel 105 homogenized forabout 10 minutes. The mixture in the pre-mixing vessel 105 is thenpumped through the peristaltic pump 155 to the mixing vessel 160 (block2240), for example at a rate of 500 mL/min. The pre-mixing vessel 105 isthen rinsed with ethyl alcohol for about 5 minutes at a homogenizationrate of 4000 revolutions per minute (blocks 2260 and 2270). After 5minutes, the rate of the homogenizer 110 is reduced to 2500 revolutionsper minute and the contents transferred to the mixing vessel 160 throughthe peristaltic pump 155, for example at a rate of 500 mL/min (block2270). The combined contents of the mixing vessel 160 are mixed at amixing rate of about 45 RPM for about an additional 90 minutes (block2280). The mixed material may then be stored in any suitable containerthat avoids contact with moisture or may be pumped to a packaging systemas described herein.

As will be appreciated by one of skill in the art in light of thepresent disclosure, the step/operations illustrated in may be carriedout of sequence or concurrently or steps/operations may be combinedwhile still falling within the scope of the present inventive concept.For example, if multiple rotor/stator homogenizing vessels are utilizedwhat is illustrated in the Figures as a sequence of steps may be carriedout simultaneously. Furthermore, operations which are illustrated asbatch operations could be modified to provide continuous processoperations. Accordingly, the flowchart illustrations should not beconstrued as limiting the scope of the present invention.

FIG. 4A is a cross section drawing of a pre-mixing vessel 105 suitablefor use in the system 100. FIG. 4B is a drawing of the lid and FIG. 4Cis a drawing of the rotar-stator for the homogenizer 110. As seen inFIG. 4A, the pre-mixing vessel 105 includes a homogenizer vessel 110with a jacket 120 for passing heating/cooling media around thehomogenizer vessel 110. The heating/cooling media enters and exits thejacket 120 through the inlet/outlet ports 111, 112. The homogenizervessel 110 has a drain port 113 with a drain valve 114 that is a zeroclearance valve so as to not leave any dead space in the homogenizervessel 110 where ingredients could collect. Furthermore, the drain port113 and drain valve 114 are large enough to provide for the rapidremoval of material from the pre-mixing vessel 105 and so that it willnot clog with the gel. The pre-mixing vessel 105 may be made of anysuitable material but, in some embodiments, it is made of glass.

FIG. 4B illustrates the lid 115 of the pre-mixing vessel 105. The lid115 includes three openings 116, 117, 118 that may be sealed and allowaccess to the homogenizer vessel 110. The largest opening 116 is for therotor/stator 119 of the homogenizer. The middle opening 117 is for theinert gas. The smallest opening is for a temperature probe 118. Therotor/stator 119 illustrated in FIG. 4C allows for a seal where theshaft goes through the lid 115.

In a particular embodiment, the dimension PN is 170 mm, L1 is 200 mm, IDis 170 mm and L2 is 300 mm. The drain valve 114 is 20 mm. The opening116 is 30 mm in diameter, the opening 117 is 20 mm in diameter and theopening 118 is 8 mm in diameter. The rotor/stator 119 is model numberT50 from IKA-Werk. The use of the rotor/stator 119 allows for theplacement of the opening 116 centered or off center in the lid 115.

While embodiments of the present inventive subject matter have beendescribed with reference to particular configurations of a processingsystem and specific processing conditions variations to these systemsand conditions may be made while still falling within the scope of thepresent inventive subject matter. For example, the processing system mayinclude solid powder metering systems for raw materials, such as theKlucel and the API. Similarly, additional systems could be incorporatedto prevent oxidation of the materials. For example, processing could becarried out under vacuum to remove oxygen and carbon dioxide, solutionscould be degassed via vacuum or bubbling, the materials could beprotected from light (e.g. if a nitrosothiol NO releasing donor isutilized) or light could be used to either photo-age the topical gel oreliminate photodecomposition of the topical gel or its components.

Packaging and Delivery of Topical Compositions

Returning to FIG. 1, also illustrated is a packaging system 300. As seenin FIG. 1, the packaging system 300 includes a hopper 310 for receivingthe product to be packaged and a tube filler 320, such as an Arencomatic1000 from Norden, for taking the product from the hopper 310, dispensingthe product into a tube and sealing the tube. Argon gas is provided inthe hopper 310 and to the tube filler 320 with the rate of flow beingcontrolled by the tube filler 320 or a flow meter 330. A blanket ofargon gas is provided in the hopper 310 and the hopper 310 filled fromthe bottom so that the product is not exposed to moisture in the air.The tube filler 320 fills preformed tubes with argon gas and then fillsthe tubes from the bottom to displace the gas and, thereby, reduce thelikelihood that the product will be exposed to moisture.

In some embodiments, a nitric oxide-releasing topical therapeutic may beprovided in sealed single dose tubes (e.g., 2 g tubes) that are oxygenimpermeable to prevent moisture from reaching the topical therapeuticand causing release of the nitric oxide. The patient would then puncturethe tube at the time of application and apply the product to their skin.The product could also be refrigerated until use to thereby furtherreduce the likelihood of premature release of the nitric oxide.

In addition to being oxygen impermeable, the coatings and adhesives ofthe tubes should not contaminate the topical gel in the tubes. Inparticular, the adhesive at the crimped end of the tube should notcontain a cold seal. Adhesives such as Darex™ and related syntheticrubber and latex compounds, which could have an adverse reaction andsubsequently could contaminate the topical gel such as initiatingrelease of nitric oxide, changing the color of the gel or otherundesirable result. In some embodiments, packaged nitric oxide-releasingtopical gels may be provided with a shelf life of at least about oneweek.

In further embodiments, the packaged nitric oxide-releasing topical gelshave a shelf life of at least about four weeks, at least about 12 weeks,at least about 26 weeks, or at least about 52 weeks. In still furtherembodiments, the packaged nitric oxide-releasing topical gels have ashelf life of from at least 12 to at least 104 weeks, or any rangetherein. As used herein, the term “shelf life” refers to the length oftime a product (i.e., a topical gel of the present invention) maintainsthe ability to release a therapeutically effective amount of nitricoxide in an unopened package stored under recommended storageconditions. The shelf life may, for example, be evidenced by the “useby” or “best if used by” date for the product, the manufacturer'sexpiration date of the product and/or the actual product characteristicsafter the specified period of time. Accordingly, the term “shelf life”as used herein should be construed as including both an “actual” shelflife of the product and a “predicted” shelf life of the product unlessstated otherwise. As one skilled in the art will recognize, the rate ofrelease of nitric oxide in a topical gel under packaged and/or storedconditions may be different (i e, faster or slower) than the rate ofrelease of nitric oxide when the topical gel is in use. In certainembodiments, the rate of release of nitric oxide may be faster when atopical gel is in use compared to the rate of release of nitric oxidewhen the topical gel was packaged and/or stored.

In some embodiments, the shelf life of the product is the time that theproduct maintains the ability to release at least 50% of the initialamount of nitric oxide that the product may release when packaged. Infurther embodiments, the shelf life of the product is the time that theproduct maintains the ability to release at least 70%, at least 80%, atleast 90%, at least 95%, or at least 98% of the initial amount of nitricoxide that the product may release when packaged. In some embodiments,the recommended storage conditions are room temperature. In someembodiments, the recommended storage conditions are refrigerated storageconditions. In particular embodiments, the refrigerated storageconditions are between 1° C.-12° C., or any range therein.

Further embodiments may provide packaged nitric oxide-releasing topicalgels that have a useful life of at least about 7 days after opening thepackage. In further embodiments, the useful life is at least about 30days, at least about 60 days or at least about 90 days. In still furtherembodiments, the packaged nitric oxide-releasing topical gels have auseful life of from at least about 60 days to at least about 730 days.As used herein, the term “useful life” refers to the length of time thatthe product maintains the ability to release a therapeutically effectiveamount of nitric oxide from an opened packaged when applied asrecommended and when stored under recommended storage conditions. Theuseful life may, for example, be evidenced by the manufacturer'srecommended time to dispose of the product after opening or measurementsof the products characteristics after opening.

Accordingly, the term “useful life” as used herein should be construedas including both an “actual” useful life of the product or a“predicted” useful life of the product unless stated otherwise. In someembodiments, the useful life of the product is the time that the productmaintains the ability to release at least 50% of the initial amountnitric oxide that the product may release when the package is opened. Infurther embodiments, the shelf life of the product is the time that theproduct maintains the ability to release at least 70%, at least 80%, atleast 90%, at least 95%, or at least 98% of the initial amount nitricoxide that the product may release when the package is opened. In someembodiments, the recommended storage conditions after opening are roomtemperature. In particular embodiments, the recommended storageconditions after opening are refrigerated conditions.

Methods of Using Anhydrous Topical Compositions

A composition described herein may be used to treat a dermatologicalailment. For example, a composition of the present invention may be usedto treat acne. In certain embodiments, the composition comprises atleast one API, such as a NO-releasing API. In other embodiments, thecomposition comprises a vehicle and thus does not comprise an API. Acomposition described herein may be applied to the skin at aconcentration sufficient to treat a dermatological ailment such as acne.As such, in some embodiments, a topical composition described herein maydecrease, eliminate or prevent acne, decrease abnormal keratinization,reduce microbial colonization, reduce inflammation and/or decrease sebumproduction. Decrease of acne may be detected by a visual reduction inthe amount or severity of the acne and/or by decrease in pain ordiscomfort associated with the acne, as identified by the subject.

In particular embodiments of the present invention, a topicalcomposition described herein is non-drying and/or non-irritating to theskin of a subject. Dryness and/or irritation of the skin may be detectedvisually and/or as identified by a subject.

In some embodiments, an anhydrous topical composition according toembodiments of the invention is applied topically to the skin of thesubject. Any portion of the subject's skin may be treated. However, insome embodiments, the subject's face is treated by a method describedherein. Furthermore, in some embodiments, the subject's trunk is treatedby a method described herein. The subject may have acne or apharmaceutical composition described herein may be used to prevent acnefrom forming.

Subjects suitable to be treated with a method of the invention include,but are not limited to, avian and mammalian subjects. Mammals of thepresent invention include, but are not limited to, canines, felines,bovines, caprines, equines, ovines, porcines, rodents (e.g. rats andmice), lagomorphs, primates, humans, and the like, and mammals in utero.Any mammalian subject in need of being treated according to the presentinvention is suitable. Human subjects of both genders and at any stageof development (i.e., neonate, infant, juvenile, adolescent, adult) maybe treated according to the present invention.

Illustrative avians according to the present invention include chickens,ducks, turkeys, geese, quail, pheasant, ratites (e.g., ostrich) anddomesticated birds (e.g., parrots and canaries), and birds in ovo.

The invention may also be carried out on animal subjects, particularlymammalian subjects such as mice, rats, dogs, cats, livestock and horsesfor veterinary purposes, and for drug screening and drug developmentpurposes.

In some embodiments, a method of treating acne, decreasing abnormalkeratinization, reducing microbial colonization, reducing inflammationand/or decreasing sebum production may include using a method describedherein in combination with another therapeutic regimen and/or incombination with another API, such as those that have antimicrobial,anti-inflammatory, pain-relieving, immunosuppressant, vasodilatingproperties, and/or anti-acne properties. For example, other anti-acneagents such as retenoids, may be used in conjunction (prior,concurrently or after) with the application of the nitric oxide. Assuch, in some embodiments of the invention, a patient may be treatedwith nitric oxide, at least one NO releasing compound and/or acomposition described herein in combination with an additionaltherapeutic agent when the additional therapeutic agent is not in thesame composition. For example, in some embodiments, an additionaltherapeutic agent may be administered (e.g., topically, systemically,parenterally, orally, buccally, subcutaneously, via inhalation,intratracheally, surgically, transdermally, etc.), either concurrentlyand/or sequentially with application of nitric oxide, at least onenitric oxide source and/or a pharmaceutically acceptable compositionthat includes at least one nitric oxide source.

In some embodiments of the invention, a pharmaceutically acceptablecomposition described herein may be administered to the skin via spraydelivery. A non-aqueous delivery propellant may be used for watersensitive NO-releasing compounds including diazeniumdiolate-modifiedcompounds. Further, in some embodiments, particular components of themedicaments may be separated at some point prior to application of themedicament. For example, a water reactive NO-releasing compound may bestored separately from an aqueous component or propellant untilapplication (e.g., via spraying or applying a gel). In some embodiments,the NO-releasing compounds may be combined with an aqueous constituentprior to application or the NO-releasing compounds and an aqueousconstituent may be applied to the skin sequentially.

While some embodiments of the present inventive concept may be suitablefor treating acne so as to moderate production of sebum, moderateabnormal keratinization, reduce inflammation and/or reduce microbialcolonization, some embodiments may promote healing of a wound associatedwith the dermatological condition, affect a biofilm associated with thedermatological condition and/or reduce scarring from a wound associatedwith the dermatological condition. Additionally, embodiments of thepresent inventive concept may also treat the dermatological conditionwithout significant side effects.

Accordingly, particular embodiments of the present inventive conceptprovide a method of treating a dermatological condition by topicallyadministering a nitric oxide releasing topical composition in an amounteffective to actively promote healing of a wound associated with thedermatological condition. Some embodiments provide a method of treatinga dermatological condition by topically administering a nitric oxidereleasing composition in an amount effective to affect a biofilmassociated with the dermatological condition. Still further embodimentsprovide a method of treating a dermatological condition by topicallyadministering a nitric oxide releasing composition in an amounteffective to reduce scarring from a wound associated with thedermatological condition. Some embodiments provide a method of treatinga dermatological condition by topically administering a nitric oxidereleasing composition in an amount effective to treat the dermatologicalcondition without significant side effects.

Wound healing may be effected through prolonged low concentrations ofnitric oxide administration whereby nitric oxide acts as a signalingmolecule in a number of wound healing cascades. An intermediateNO-releasing composition may mitigate the inflammatory phase. Highlevels of NO-released may be necessary to effect antimicrobial activity,using the rapid burst of nitric oxide to kill microorganisms throughoxidative/nitrosative intermediates. In some embodiments, provided is akit for the topical treatment of dermatological ailments that includesat least one composition that includes at least one viscosity increasingagent, at least one organic solvent, at least one humectant and at leastone water repellant. In some embodiments, a kit includes a firstcomposition that releases low concentrations of nitric oxide; and asecond composition that releases a higher level of nitric oxide.Additional APIs may be included in some or all of the compositions toincrease efficacy.

While embodiments of the present invention have been described asproviding a topical composition that includes an API, in someembodiments, a topical composition according to the present inventionmay be used to treat a condition and, in particular, a dermatologicalcondition as described herein, such as acne, without the inclusion of anAPI. In such embodiments of the present invention, the “vehicle” may beused to treat the dermatological condition. Accordingly, therapeuticapplications of a topical composition of the present invention includeapplication of a topical composition with and/or without an API includedin the topical composition.

EXAMPLES Example 1 Examples of Topical Gels

TABLE 1 Nitricil ™ Gel, 0.2% Quality Component Supplier StandardFunction % w/w Ethyl alcohol, Koptec USP Solvent 85.3 anhydrous, 200proof, or BDH USP or Isopropyl alcohol, USP Hexylene glycol, Fluka/Reagent Co-solvent/ 10.0 99% Sigma Humectant Cyclomethicone, NF Dow NFWater- 2.5 ST-Cyclomethicone-5 Corning repelling agent HydroxypropylAshland NF Viscosity- 2.0 cellulose, NF modifying agent Klucel MF PharmNitricil ™ Novan N/A API 0.2 Total 100.0

TABLE 2 Nitricil ™ Gel, 0.5% Quality Component Supplier StandardFunction % w/w Ethyl alcohol, Koptec USP Solvent 85.0 anhydrous, 200proof, or BDH USP or Isopropyl alcohol, USP Hexylene glycol, Fluka/Reagent Co-solvent/ 10.0 99% Sigma Humectant Cyclomethicone, NF Dow NFWater- 2.5 ST-Cyclomethicone-5 Corning repelling agent HydroxypropylAshland NF Viscosity- 2.0 cellulose, NF modifying agent Klucel MF PharmNitricil ™ Novan N/A API 0.5 Total 100.0

TABLE 3 Nitricil ™ Gel, 1% Quality Component Supplier Standard Function% w/w Ethyl alcohol, Koptec USP Solvent 84.5 anhydrous, 200 proof, orBDH USP or Isopropyl alcohol, USP Hexylene glycol, Fluka/ ReagentCo-solvent/ 10.0 99% Sigma Humectant Cyclomethicone, NF Dow NF Water-2.5 ST-Cyclomethicone-5 Corning repelling agent Hydroxypropyl Ashland NFViscosity- 2.0 cellulose, NF modifying agent Klucel MF Pharm Nitricil ™Novan N/A API 1.0 Total 100.0

TABLE 4 Nitricil ™ Gel, 2% Quality Component Supplier Standard Function% w/w Ethyl alcohol, anhydrous, Koptec USP Solvent 83.5 200 proof, USPor or BDH Isopropyl alcohol, USP Hexylene glycol, 99% Fluka/ ReagentCo-solvent/ 10.0 Sigma Humectant Cyclomethicone, NF Dow NFWater-repelling 2.5 ST-Cyclomethicone-5 Corning agent Hydroxypropylcellulose, Ashland NF Viscosity- 2.0 NF Klucel MF Pharm modifying agentNitricil ™ Novan N/A API 2.0 Total 100.0

TABLE 5 Nitricil ™ Gel, 3% Quality Component Supplier Standard Function% w/w Ethyl alcohol, anhydrous, Koptec USP Solvent 82.5 200 proof, USPor or BDH Isopropyl alcohol, USP Hexylene glycol, 99% Fluka/ ReagentCo-solvent/ 10.0 Sigma Humectant Nitricil ™ Novan N/A API 3.0Cyclomethicone, NF Dow NF Water-repelling 2.5 ST-Cyclomethicone-5Corning agent Hydroxypropyl cellulose, Ashland NF Viscosity- 2.0 NFKlucel MF Pharm modifying agent Total 100.0

TABLE 6 Nitricil ™ Gel, 4% Quality Component Supplier Standard Function% w/w Ethyl alcohol, anhydrous, Koptec USP Solvent 81.5 200 proof, USPor or BDH Isopropyl alcohol, USP Hexylene glycol, 99% Fluka/ ReagentCo-solvent/ 10.0 Sigma Humectant Nitricil ™ Novan N/A API 4.0Cyclomethicone, NF Dow NF Water-repelling 2.5 ST-Cyclomethicone-5Corning agent Hydroxypropyl cellulose, Ashland NF Viscosity- 2.0 NFKlucel MF Pharm modifying agent Total 100.0

TABLE 7 Nitricil ™ Gel, 5% Quality Component Supplier Standard Function% w/w Ethyl alcohol, anhydrous, Koptec USP Solvent 80.5 200 proof, USPor or BDH Isopropyl alcohol, USP Hexylene glycol, 99% Fluka/ ReagentCo-solvent/ 10.0 Sigma Humectant Nitricil ™ Novan N/A API 5.0Cyclomethicone, NF Dow NF Water-repelling 2.5 ST-Cyclomethicone-5Corning agent Hydroxypropyl cellulose, Ashland NF Viscosity- 2.0 NFKlucel MF Pharm modifying agent Total 100.0

TABLE 8 Nitricil ™ Gel, 6% Quality Component Supplier Standard Function% w/w Isopropyl alcohol, USP Koptec USP Solvent 80.5 or BDH Hexyleneglycol, 99% Fluka/ Reagent Co-solvent/ 10.0 Sigma Humectant Nitricil ™Novan N/A API 6.0 Cyclomethicone, NF Dow NF Water-repelling 2.5ST-Cyclomethicone-5 Corning agent Hydroxypropyl cellulose, Ashland NFViscosity- 1.0 NF Klucel MF Pharm modifying agent Total 100.0

TABLE 9 Nitricil ™ Gel, 12% Quality Component Supplier Standard Function% w/w Isopropyl alcohol, USP Koptec USP Solvent 74.5 or BDH Hexyleneglycol, 99% Fluka/ Reagent Co-solvent/ 10.0 Sigma Humectant Nitricil ™Novan N/A API 12.0 Cyclomethicone, NF Dow NF Water-repelling 2.5ST-Cyclomethicone-5 Corning agent Hydroxypropyl cellulose, Ashland NFViscosity- 1.0 NF Klucel MF Pharm modifying agent Total 100.0

TABLE 10 Nitricil ™ Gel, 20% Quality Component Supplier StandardFunction % w/w Isopropyl alcohol, USP Koptec USP Solvent 66.5 or BDHHexylene glycol, 99% Fluka/ Reagent Co-solvent/ 10.0 Sigma HumectantNitricil ™ Novan N/A API 20 Cyclomethicone, NF Dow NF Water-repelling2.5 ST-Cyclomethicone-5 Corning agent Hydroxypropyl cellulose, AshlandNF Viscosity- 1.0 NF Klucel MF Pharm modifying agent Total 100.0

TABLE 11 Nitricil ™ Gel, 30% Quality Component Supplier StandardFunction % w/w Isopropyl alcohol, USP Koptec USP Solvent 56.5 or BDHHexylene glycol, 99% Fluka/ Reagent Co-solvent/ 10.0 Sigma HumectantNitricil ™ Novan N/A API 30.0 Cyclomethicone, NF Dow NF Water-repelling2.5 ST-Cyclomethicone-5 Corning agent Hydroxypropyl cellulose, AshlandNF Viscosity- 1.0 NF Klucel MF Pharm modifying agent Total 100.0

TABLE 12 Placebo Gel Quality Component Supplier Standard Function % w/wEthyl alcohol, anhydrous, Koptec USP Solvent 85.5 200 proof USP or orBDH Isopropyl alcohol, USP Hexylene glycol, 99% Fluka/ ReagentCo-solvent/ 10.0 Sigma Humectant Cyclomethicone, NF Dow NFWater-repelling 2.5 ST-Cyclomethicone-5 Corning agent Hydroxypropylcellulose, Ashland NF Viscosity- 2.0 NF Klucel MF Pharm modifying agentTotal 100.0

Example 2 NO-Releasing Topical Gels P. Acne Time Kill

A 2% API loaded anhydrous gel of a specific embodiment described herein,wherein the API is a nitric oxide releasing macromolecule, was testedfor efficacy in the reduction of P. acnes, using ASTM E-2315. In thistest, Propionibacterium acnes ATCC 6919 (10⁶ CFU/ml) were incubated with250 mg of gel vehicle or 250 mg of active gel containing Nitricil™ (4mg/ml) in 37° C. phosphate buffered saline (PBS). After 30 minutes, 2hours, and 4 hours of incubation, aliquots from each tube were removed,neutralized with letheen broth, serially diluted, and plated to obtainviable colony counts.

TABLE 13 Control Blank Time, hrs Time, hrs Control 0 0.5 1 2 Control 00.5 1 2 1 2.10E+06 1.60E+06 2.00E+06 1.60E+06 1 .32 .20 .30 .20 22.20E+06 1.60E+06 1.80E+06 1.90E+06 2 .34 .20 .26 .28 Average 2.15E+061.60E+06 1.90E+06 1.75E+06 Average .33 .20 .28 .24 STDERR 3.54E+040.00E+00 7.07E+04 1.06E+05 STDERR .01 .00 .02 .03 % 0 26 12 19 Log .00.13 .05 .09 Reduction Reduction vs ctrl vs Ctrl

TABLE 14 Gel vehicle by itself Gel Time, hrs Gel Time, hrs Vehicle 0 0.51 2 Vehicle 0 0.5 1 2 1 2.30E+06 1.80E+06 1.80E+06 2.90E+05 1 .36 .26.26 .46 2 2.10E+06 2.00E+06 2.10E+06 3.30E+05 2 .32 .30 .32 .52 Average2.20E+06 1.90E+06 1.95E+06 3.10E+05 Average .34 .28 .29 .49 STDERR7.07E+04 7.07E+04 1.06E+05 1.41E+04 STDERR .01 .02 .02 .02 % −2 12 9 86Log 0.01 .05 .04 .84 Reduction Reduction vs ctrl vs Ctrl

TABLE 15 Gel vehicle with Nitricil ™ 2% 2% Nitricil ™ Nitricil ™ GelTime, hrs Gel Time, hrs 11-9-37 0 0.5 1 2 11-9-37 0 0.5 1 2 1 1.90E+062.10E+03 1.00E+01 1.00E+01 1 .28 .32 .00 .00 2 1.80E+06 2.10E+031.00E+01 1.00E+01 2 .26 .32 .00 .00 Average 1.85E+06 2.10E+03 1.00E+011.00E+01 Average .27 .32 .00 .00 STDERR 3.54E+04 0.00E+00 0.00E+000.00E+00 STDERR .01 .00 .00 .00 % 14 99.90 99.9995 99.9995 Log .07 01.33 .33 Reduction Reduction vs ctrl vs Ctrl

TABLE 16 Summary of Data % Time, hrs Log Time, hrs Reduction 0 0.5 1 2Reduction 0 0.5 1 2 Control 6 12 19 Control .00 .13 .05 .09 Gel Vehicle2 2 9 86 Gel Vehicle 0.01 .05 .04 .84 2% 4 9.90 >99.999 >99.999 2% .07.01 5.33 5.33 Nitricil ™ Nitricil ™ Gel Gel

The data in Tables 13-16 is also shown graphically in FIG. 5. As can beseen from Table 16 and FIG. 5, a NO-releasing composition according toan embodiment of the invention can be effective to kill P. acnes.

Example 3

The NO release profile for 2% Nitricil™ in anhydrous gel, as tested inExample 1 and made by the process described herein, is shown in FIG. 6.At a pH of 6.0 that may be encountered in the stratum corneum, themaximum flux of nitric oxide exceeded 0.5 nmol NO per mg gel per secondand a half-life of about 160 seconds.

Example 4

Stability data for 2% Nitricil™ in anhydrous gel, as tested in Example 1and made by the process described herein, are shown in FIG. 7

Example 5 Macromolecular Nitric Oxide Releasing Compound

A nitric oxide releasing macromolecular compound comprising MAP3 wasfabricated as described in United States Patent Application PublicationNo. 2009/0214618 and in PCT Patent Application Number PCT/US12/22048,filed Jan. 20, 2012, entitled “Temperature Controlled Sol-GelCo-Condensation,” the disclosures of which are incorporated herein byreference as if set forth in their entirety. The resultingmacromolecular particles were ball milled to provide an average particlesize of from 8 to 10 μm to provide an active pharmaceutical ingredient(API).

Example 6 Topical Gel with Active Pharmaceutical Ingredient

Topical gel incorporating the API of Example 5 (“Active”) was fabricatedas described in U.S. Provisional Patent Application Ser. No. 61/504,628entitled “Topical Compositions and Methods of Using Same to Treat Acne,”and U.S. Provisional Patent Application Ser. No. 61/504,626, entitled“Methods of Manufacturing Topical Compositions and Apparatus for Same,”both of which were filed Jul. 5, 2011 and are incorporated herein as ifset forth in their entirety. In particular, the Active had theformulation of:

TABLE 17 Active Gel Quality Component Standard Function % w/w Ethylalcohol, anhydrous, 200 USP Solvent 83.5 proof, USP Hexylene glycol, 99%Reagent Co-solvent/ 10.0 Humectant Cyclomethicone, NF NF Water-repelling2.5 ST-Cyclomethicone-5 agent Hydroxypropyl cellulose, NF NF Viscosity-2.0 Klucel MF Pharm modifying agent Nitricil ™ N/A API 2.0

Example 7 Topical Gel with Masking Agent

Topical gel incorporating titanium dioxide as a masking agent(“Vehicle”) was fabricated as described in U.S. Provisional PatentApplication Ser. No. 61/504,628 entitled “Topical Compositions andMethods of Using Same to Treat Acne,” and U.S. Provisional PatentApplication Ser. No. 61/504,626, entitled “Methods of ManufacturingTopical Compositions and Apparatus for Same,” both of which were filedJul. 5, 2011 and are incorporated herein as if set forth in theirentirety. In particular, the Vehicle had the formulation of:

TABLE 18 Vehicle Gel Quality Component Standard Function % w/w Ethylalcohol, anhydrous, 200 USP Solvent 85.45 proof, USP Hexylene glycol,99% Reagent Co-solvent/ 10.0 Humectant Cyclomethicone, NF NFWater-repelling 2.5 ST-Cyclomethicone-5 agent Hydroxypropyl cellulose,NF NF Viscosity- 2.0 Klucel MF Pharm modifying agent Titanium dioxideN/A 0.05

All clinical trial materials were packaged in single use 2 g tubes on anArencomatic 1000 automated packaging system from Norden. The 2 g tubeswere stored at 5° C. until dispensing to trial subjects. The tubespackaged into 2 week kits.

Example 8 Clinical Study Protocol

A single-center, double-blind, randomized, vehicle-controlled, parallelgroup study was conducted in 70 intent-to-treat (ITT) subjects withmoderate to severe acne vulgaris. The study included male and femalesubjects between the ages of 12 and 40 (inclusive) with moderate tosevere acne vulgaris, defined as at least 20 but no more than 40inflammatory lesions (papules and pustules), 20 to 60 non-inflammatorylesions (open and closed comedones), no more than 2 nodules, and anInvestigator Global severity Assessment of 3 or 4. Subjects whosatisfied the entry criteria at the Baseline visit were randomized toeither Active or Vehicle in a 1:1 ratio. Tolerability and safetyassessments included cutaneous tolerability evaluation, physical exams,collection of vital signs, blood pressure monitoring, urine pregnancytests (UPTs), blood methemoglobin monitoring and adverse eventcollection. Efficacy assessments included inflammatory andnoninflammatory lesion counts and an Investigator global severityassessment (IGA). Subjects returned for post-baseline evaluation atWeeks 2, 4, 6 and 8. Of the 70 ITT subjects enrolled in the study, 60subjects completed the study per the study protocol. The subjects wereprovided kits with a two week supply of the 2 g tubes at each visit andinstructed to maintain the tubes under refrigerated conditions untiluse.

Example 9 Tolerability and Safety Results

Cutaneous tolerability results of the study are shown in Table 19 forthe Active and Table 20 for the Vehicle.

TABLE 19 Summary of Cutaneous Tolerability at Baseline and Week 8 -Active Baseline Pre- Application Maximum Severity Week 8 Mild Mod SevereMild Mod Severe Mild Mod Severe Erythema 1 0 0 4 2 0 1 0 0  (2.9) % (12.1) % (5.7%) (3.0) % Scaling 0 0 0 2 2 0 0 1 0 (5.7%)  (5.7%) (3.0)% Dryness 0 0 0 0 0 0 0 0 0 Itching 3 2 0 4 2 0 2 2 0 (8.6%) (5.7%)(11.8%) (6.1%) (6.1%)  (6.1%)  Burning/ 0 0 0 1 1 0 1 0 0 Stinging(3.0)%  (3.0%) (3.0) %

TABLE 20 Summary of Cutaneous Tolerability at Baseline and Week 8 -Vehicle Baseline Maximum End of Treatment Pre-Application Severity (Wk8) Mild Mod Severe Mild Mod Severe Mild Mod Severe Erythema 1 0 0 2 0 00 0 0 (2.9)%  (5.7%) Scaling 0 0 0 1 0 0 0 0 0 (2.9%) Dryness 0 0 0 0 00 0 0 0 Itching 6 1 0 1 0 0 0 0 0 (17.1%) (2.9%) (2.9%) Burning/ 0 0 0 10 0 1 0 0 Stinging (2.9%) (3.6%)

Methemoglobin results were obtained by pulse co-oximeter from Masimo.The results at baseline, 2 weeks and 8 weeks are shown in Table 21.

TABLE 21 Methemoglobin Levels as Measured by Pulse Co-Oximetry Baseline2 Weeks 8 Weeks Active N 35 35 34 Mean 0.9 1.1 0.9 STD 0.3 0.3 0.3Vehicle N 35 34 31 Mean 0.7 1.0 0.8 STD 0.3 0.3 0.4

As seen in Table 21, the methemoglobin levels for Active and Vehiclesubjects were will within acceptable ranges and did not differsignificantly.

Example 10 Shelf Life

Tubes of the clinical supplies were maintained under controlledconditions at 5° C. and periodically evaluated for nitric oxide content.Tubes were also maintained at a controlled storage facility at 5° C. bythe packager/labeler that prepared the kits and evaluated for nitricoxide content. Unused materials distributed to and retrieved from theclinical study site in the Dominican Republic were also evaluated fornitric oxide content. The results of the evaluation for remaining nitricoxide are shown in Table 22.

TABLE 22 Shelf Life Stability of 2% Topical Gel in 2 gram Tubes ActiveGel % Recovery vs Initial Relative Std 5° C. NO Loading Dev 6 monthStability @ Storage 96.6% N/A Facility (n = 2) Retained ClinicalMaterials  100% 0.5% (n = 3) Material Retuned from Clinical 93.1% 4.6%Trial (n = 4)

Example 11 NO recovery over time

Tubes of a 2% anhydrous gel were stored at 5° C., 25° C./60% relativehumidity (R.H.), 30° C./65% R.H., and 40° C./75% R.H. At 2, 4, 8, 12 and16 weeks, the gel in the tubes was evaluated for nitric oxide content.FIG. 8 shows the nitric oxide recovery for the 2% anhydrous gels in 2gram tubes over a time. The results of the stability study are providedin Table 23.

TABLE 23 Stability of 2% anhydrous gel at various temperatures over 26weeks % NO % NO Temperature/ % NO % NO % NO Recovered Recovered RelativeRecovered Recovered Recovered at at Humidity at Week 2 at Week 4 at Week8 Week 12 Week 26 5° C. 98.27586 100 86.2069 94.82759 96.55172 25°C./60% 81.03448 74.13793 58.62069 65.51724 53.44828 R.H. 30° C./65%74.13793 67.24138 53.44828 48.27586 25.86207 R.H. 40° C./75% 50 37.9310320.68966 13.7931 6.896552 R.H.

Example 12 Activity of Vehicle for P. Acnes in Healthy Patients

Topical Gel Vehicle with titanium dioxide was produced as described inExample 7. Topical Gel Vehicle without titanium dioxide was produced asdescribed in Example 7 except the titanium dioxide was replaced withadditional ethyl alcohol. These formulations were evaluated to determineif the Vehicle in the clinical study had antimicrobial activity. Avehicle with antimicrobial activity could mask the effectiveness of anactive ingredient in a clinical study.

Topical Gel Vehicle with titanium dioxide and Topical Gel Vehiclewithout titanium dioxide were evaluated in vivo to determine the effectson P. acnes and cutaneous tolerability. Thirty-one healthy adult malesand females 18 years of age and older were enrolled in the study aftermeeting eligibility requirements at screening, 7-10 days prior tobaseline. Subjects were carefully screened to ensure that none wereusing any prohibited topical or systemic antibiotics within 4 weeksprior to enrollment. The panelists were instructed not to use anyantimicrobial topical products (e.g., soaps, medicated shampoos, acnepreparations) during the study. The volunteers were selected for thestudy based on a high degree of fluorescence of the facial skin under aWood's lamp indicating the presence of high levels of P. acnes. BaselineP. acnes counts were at least 10,000 colonies per cm² on the facialskin.

Subjects were treated for four weeks with either Topical Gel Vehiclewith titanium dioxide or Topical Gel Vehicle without titanium dioxide.Each volunteer was treated once daily on the central forehead undersupervision by a technician at the site beginning on the Thursdayfollowing acceptance into the study. Topical Gel Vehicle was alsoapplied by the subjects (unsupervised) at home once daily on Saturdaysand Sundays. At each visit, a sufficient amount of the test product(pea-sized amount) was applied and rubbed in for about 30 seconds.

Quantitative bacteriologic cultures were obtained from the test site atBaseline (0) and at Weeks 2 and 4. During the study, cutaneoustolerability assessments were made at Baseline and Weeks 1, 2, 3 and 4.Each of the local signs and symptoms, erythema, dryness, scaling,stinging/burning and itching were assessed on a four point scale where0=none, 1=mild, 2=moderate, and 3=severe. Thirty-one patients wereenrolled and 30 patients completed the study. One patient wasdiscontinued prematurely by the Investigator for non-compliance.

Table 24 includes the P. acnes counts for the 4 week study. As seen inTable 24, neither of the formulations tested achieved significantreductions in P. acnes.

TABLE 24 Total P. acnes Total P. acnes (Log/cm²) Topical Gel VehicleTopical Gel Vehicle without titanium dioxide with titanium dioxide Mean± Std over Mean ± Std over Baseline  6.75± N/A 6.72± N/A Week 2  6.66±0.0180 6.69± 0.0000 Week 4  6.64± 0.0037 6.67± 0.0000 Net Change fromBaseline Week 2 −0.08 ± 0.12 N/A −0.05±  N/A Week 4 −0.11 ± 0.12 N/A−0.07±  N/A

With regard to the cutaneous tolerability assessment, there was noerythema, scaling or itching observed/reported during the study witheither formulation. There was only one report of mild (score=1)burning/stinging at Week 1 with the topical gel without titanium dioxideand one report of mild (score=1) dryness with the topical gel withtitanium dioxide at Week 4.

Many modifications and other embodiments of the inventions set forthherein will come to mind to one skilled in the art to which theseinventions pertain having the benefit of the teachings presented in theforegoing descriptions and the associated drawings. Therefore, it is tobe understood that the inventions are not to be limited to the specificembodiments disclosed and that modifications and other embodiments areintended to be included within the scope of the appended claims.Although specific terms are employed herein, they are used in a genericand descriptive sense only and not for purposes of limitation.

1. An anhydrous topical composition consisting essentially of: at leastone viscosity increasing agent at a concentration in a range of 0.5% to30% by weight of the anhydrous topical composition, at least one organicsolvent at a concentration in a range of 50% to 90% by weight of theanhydrous topical composition, at least one humectant at a concentrationin a range of 2% to 20% by weight of the anhydrous topical composition,at least one water repellent at a, concentration in a range of 0.5% to15% by weight the anhydrous topical composition, and at least oneanti-acne agent.
 2. (canceled)
 3. The anhydrous topical composition ofclaim 1, wherein the at least one viscosity increasing agent compriseshydroxypropylcellulose; the at least one organic solvent comprisesethanol; the at least one humectant comprises hexylene glycol. 4.-5.(canceled)
 6. The anhydrous topical composition of claim 1, wherein theat least one viscosity increasing agent compriseshydroxypropylcellulose; the at least one organic solvent comprisesethanols the at least one humectant comprises hexylene glycol; and theat least one water repellent comprises cyclomethicone. 7.-18. (canceled)19. The anhydrous topical composition of claim 1, wherein the anhydroustopical composition is non-drying and/or non-irritating.
 20. A method oftreating a dermatological condition comprising applying the anhydroustopical composition of claim 1 to the skin of a patient to treat thedermatological condition.
 21. The method of claim 20, wherein thedermatological condition comprises acne.
 22. The anhydrous topicalcomposition of claim 1, wherein the at least one anti-acne agent isselected from the group consisting of: adapalene, azelaic acid, benzoylperoxide, clindamycin, clindamycin phosphate, doxycycline, doxycyclinemonohydrate, erythromycin, salicylic acid, retinoic acid, norgestimate,organic peroxides, isotretinoin, tretinoin, sulfacetamide sodium,tazarotene, etretinate, acetretin, and any combination thereof.
 23. Theanhydrous topical composition of claim 1, wherein the at least oneanti-acne agent is a keratolytic agent.
 24. The anhydrous topicalcomposition of claim 1, wherein the at least one anti-acne agent issalicylic acid.
 25. The anhydrous topical composition of claim 1,wherein the at least one anti-acne agent is present in the anhydroustopical composition at a concentration in a range of 0.01% to 30% byweight of the anhydrous topical composition.
 26. The anhydrous topicalcomposition of claim 1, wherein the at least one anti-acne agent ispresent in the anhydrous topical composition at a concentration in arange of 0.01% to 2% by weight of the anhydrous topical composition. 27.The anhydrous topical composition of claim 24, wherein the at least oneanti-acne agent is present in the anhydrous topical composition at aconcentration in a range of 0.01% to 2% by weight of the anhydroustopical composition.
 28. An anhydrous topical composition consistingessentially of: at least one viscosity increasing agent at aconcentration in a range of 0.5% to 30% by weight of the anhydroustopical composition; at least one organic solvent at a concentration ina range of 50% to 90% by weight of the anhydrous topical composition; atleast one humectant at a concentration in a range of 2% to 20% by weightof the anhydrous topical composition; and at least one water repellentat a concentration in a range of 0.5% to 15% by weight the anhydroustopical composition.
 29. The anhydrous topical composition of claim 28,wherein the at least one viscosity increasing agent compriseshydroxypropylcellulose; the at least one organic solvent comprisesethanol; the at least one humectant comprises hexylene glycol; and theat least one water repellent comprises cyclomethicone.
 30. A method oftreating a dermatological condition comprising applying the anhydroustopical composition of claim 28 to the skin of a patient to treat thedermatological condition.
 31. The method of claim 30, wherein thedermatological condition comprises acne.